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The interaction between 5-hydroxytryptamine and tryptophan: a serotonin receptor model
Journal of Molecular Structure (Theochem), 235 (1991) 321-342
321
Elsevier Science Publishers B.V., Amsterdam
The interaction between 5-hydroxytryptamine tryptophan: a serotonin receptor model
and
Lester A. Rubenstein”and Roman OsmarPb Departments of Physiology and Biophysics” and of Pharmucokgy’, Mount Sinai School of Medicine of the City Uniuersity of New York, New York, NY 10029 (USA)
(Received 5 November 1990)
Abstract A system consisting of serotonin f5-hydroxytryptamine, 5H’I’) and.tryptophan was used to study the process of recognition at a 5-HT receptor. The optimal geometries of interaction between 5-hy~x~ndole (5-HIND), a model ,of 5-HT, and indole, a model of tarpon, were obtained from a scan which included both eiectrostatic and dispersion energies. Recognition of 5-HIND by indole is determined by the electrostatic interaction between the components of the complex. Formation of such a complex results in electron density redistribution8 in directions both perpendicular and parallel to the planes of the interacting molecules. The nature of the charge polarization is consistent with the dominant contribution that dispersion makes to the stabilization energy of the complex. The optimal geometry of interaction between 6-HT and indole is different from the 5-HT geometry, both in the relative position of the com~nen~ of the compiex and in their orientation. This difference can be a basis for the selectivity of a receptor towards different liganda. Furthermore, the characteristica of the interaction of 5-HT with a charged imidaxolium cation, which were explored in our previous work, are different from those of the interaction of 5-HT with a neutral indole as a receptor model. In both cases, the electrostatic interaction is the o~en~tion~ determinant. With the imidaxolium cation, the main stabilization cornea from electrostatics. With the neutral indole, it comes from dispersion. Such distinctions ilhzetrate a possible basis for the specificity of ligands towards the various 5-HT receptors.
INTBODUCTION
Many ne~otr~s~t~r and drugmoleculescontain aromatic portions.Tbese portions play an important role in the selectiverecognitionof these molecules by complementaryresiduesin the receptor protein. The complementaritycan be describedin terms of the type of interaction betweenspecificportions of the receptor and the aromatic portion of the ligand. Quite often one neurotransmitter can be recognizedby different receptors. For example, serotonin (5hy~ox~~~ine, 5-HT ) is a neurotransmit~r whichacts at multiplereceptors [ 1,2], Its action on these receptors leads to different physiologicalresponses [ 3-51. The binding selectivity associated with the various receptors 0166-12~/91/$03.50
0 1991Elsevier Science Publishers B.V. All rights reserved.
[61 suggests that the mole~~&~ structures of the active sites may differ. Thus, a possible contribution to the selectivity of Merent receptors which recognize the same ligand (5-HT) may be the distinct physical nature of the interaction between ligand and receptor. In a model for a 5-HT receptor which has bsen described previously [ ‘7-10J, two elements of recognition of serotonin have been identified. The first element is an ionic interaction between the protonatedaminoethyl side-chain and a negatively charged site in the receptor. The second element is an interaction between the planar indole ring and a complement part of the receptor, which is represented by an imidazolium cation (the aromatic ring in the pro&mated form of histidine ) . Analysis of the stabilization energy of the complex between 5HT and imidazolium showed that the interaction is primarily electrostatic, and is enhanced by rn~~~ pola~~~on. Based on the changes in the charge ~tribution induced in the imidasolium cation upon its infraction with 5=HT, a proton transfer process from ~i~o~~ to a proton acceptor has been proposed f l&12 ] as a molecular m~hanism for the activation of a 5-HT receptor. The effect of a protein environment on this process has been studied in a&inidin, a protein of known structure with an appropria~ jux~~sition of the groups that form the proton transfer system proposed as an activation mechanism [ 13;14]. The indole ring in 5-HT may also interact with a neutral aromatic residue in the receptor. The electrostatic interactions between 5-HT and a neutral molecule are expected to be weaker than those with irni~~li~ cation but dispersion forces should also make an appreciable contribution to the interaction energy. The highly pol~i~ble indole ring in 5HT should give rise to favorable dispersion infractions with the pola~ble aro~tie amino acids phenyl~~ine~ tyrosine and trophy. Trophy, the most polarizabie and most polar of these three amino acids f15ftshould not only give an app~~iable dispersion interaction but should also functian as a sensitive electrostatic probe in its interactions with 5-HT. Accordingly, we have chosen to study the characteristics of the interaction between tryptophan and S-MT. Our choice of tryptophan was also guided by several experimental observations, First, close packing of indole rings is known to occur in crystals of tryptophan [X6] and 5hydroxytryptophan [ 171. Second, nuclear magnetic resonance studies of the binding of serotonin to severai oligopeptides indicate that t~toph~ undergoes a favorable infraction with 5-HT 1.IS].The bin~ng site in the peptide, co~spon~ng to residues 111-121in myelin basic protein [ 19,20], contains the essential sequence (Phe Ser Trp). A similar site in iuteinizing hormone-releasing hormone contains the sequence (Tyr Ser Trp). The binding sites in me~otropi~-stimulating hormone and adrenocorticotropic hormone each contain the sequence (Phe Arg Trp ). Thus, each of these 5HT binding sites consists of two aromatic residues fTrp, and either Phe or Tyr 1 separated by a hy~philic residue f&r or Arg ). The NMR data [ 18]
suggestthat the indole portion of 5-HT intercalatesbetween the two aromatic residues. In this study we have characterizedthe nature of the interaction between tryptophan and 5-HT and its dependence on the relative orientation of the molecules. We show that the interaction of 5-HT with a neutral molecule is different from its interactionwith a chargedcation and can serveas a basis for the selectivitybetween different 5-HT receptors. METHODS
Molecuhr geometries To reducethe size of the manta-mech~ic~ c~culations of the complexes between tryptaminesand trophy, the indole portions of these molecules wereused as models.The indoleportions of indob, 5hydroxyindole (5-HIND), 5-hydroxytryptamineand 6-hydroxyindole (6-HIND) were calculated in the Falkenberg averaged geometry [21,22]. Based upon previous findings [ 73, 5HT was calculated in the neutral form with the side-chain in the fully extended conformation. Geometriesfor benzene [ 23,241, naphthalene [ 251 and anthracene [26] were taken from crystallographicdata. Complexes between indole and !%HIND,indole and 5-HT, and indole and 6-HIND were studiedin configurationsin which the indole planes of each pair of moleculeswere kept parallel. Quantum-mechniccd
c&ulutins
Se~~cons~~nt field (SCF) c~cu~tions were done with the GAUSSI~ 80 systemof programs 1273. To reducethe numberof basis lotions, an effective core potential (ECP) [ 281 was used in conj~~ion with an LP-3G basis set [ 29 1, which was especially designed for the ECP. The interaction energy between the component molecules in a supermoleculewas decomposed by the method of Morokuma and co-workers [ 30,311 into contributionsfrom electrostatic interaction of the unperturbedmolecules (ES), mutualpolarization of the molecules (PL), exchangerepulsion(EX), chargetransfer (CT), and other residualenergycontributions (MIX). The molecularelectrostaticpotential of 5-HT was calculated from the molecular orbitals by a method described previously [ 321, and the resultswere representedas an equipotentialcontour map in a plane parallelto the indole. Electron density redistributions associated with complex formation were computed by subtracting the densities of the separated molecules from the density of the complex. The density differenceswere representedin two ways. In one method [8], density differenceswere computed in planes parallelto the indole portions by integratingthe electron density distributionover the plane.
324
The density differences were then presented as a plot of integrated density difference versus position of the plane. In a second method, density differences were calculated at points on a square grid in planes parallel to the molecules and displayed as isodensity contour maps. Calculations of stabilization energy and density difference were each corrected for basis set superposition errors (BSSE) according to the counterpoise method [ 33 1. The corrected stabilization energy, hEbB, and the corrected density difference, Ap-, are given by AE*n (corr ) = EAB-E(A+ghost)
-E(B+ghost)
Apm(corr) =PAB-P(A+ghost)
-p(B+ghost)
(I)
(2)
where AB represents the supermolecule, A and B the monomers, and a property of a given monomer is calculated using both its own basis set and the basis set of the other monomer (the ghost). Electrostatic and dispersion scan Supermolecule configurations (e.g. a complex of 5-HIND and indole) at local minima were obtained from a scan which included both electrostatic and dispersion interactions. The total interaction energy EsCANfor a given configuration of a dimer is given by E SCAN=E~~+E~I~~
(3)
where EQQdesignates the electrostatic energy, and EDIspthe dispersion energy. The electrostatic interaction energy was calculated with a monopole-monopole approximation using atomic Mulliken charges obtained from an SCF calculation with an LP-3G basis set [ 291. The electrostatic energy EQQis then given by EQQ=C
C i
j
QiQjlrG
(4)
where Qi and Qj are the Mulliken charges on the separate molecules A and B, respectively. The same sets of charges were used throughout each scan. rG is the distance between nucleus i in molecule A and nucleus j in molecule B. The dispersion energy was calculated from the interaction of bond polarizabilities according to the method of Claverie [ 341. The dispersion energy EDIsp is given by D’SP=4(UA+UB)
m ,,
(&J6
where, assuming the bonds to be axially symmetric
(5)
325
(6) VAand Unare the mean excitationenergiesof moleculesA and B, respectively. The subscriptm desi~a~s a bond in molec~a A, and the subsc~pt n a bond in molecule B. R,,= ]I$,, 1 where It,, denotes the vector joining the midpoints of bond m and bond n. A, and A, are the polar&abilitytensors of the two bonds, and T,, denotes the tensor TXn,=3(~,,X~,*)-~
(7)
where
(8) and aT is the transversecomponent of the bond polarixability.The anisotropy S of the pol~zability is given by &Z ffL-a;r
(9)
where aL is the longitudinalcomponent of the bond polarizabilityand emand e, are unit vectors in the directions of bonds m and n, respectively. Determination ofparameters for ~~u~t~~
of dispersion in~r~t~~
energy
A major cont~bution to the interaction energy between neutral molecules comes from the dispersionterm. The method of Claverie [ 341 chosen for these calculations requires the calibration of parameters for bond polarixabilities and molecular ionization potentials. Bond ~lari~bilities were calibrated by comp~ing calcula~d heats of sublimation for crystals of benzene and naphthalenewith experimentalvalues. Molecularionization potentialswere either taken from experimentor derivedfrom a linearregressionbetweenexperimental ionization potentials and calculated energiesof the highest occupied molecularorbital. For each molecule,the mean excitation energy, U, was taken to be 1.8 times the vertical ionizationpotential. This scale factor is the same as one used previously [ 351. The verticalionizationpotential of indole was taken from experimental data [SS]. Since experimentalionization potentials for 5- and 6-hydroxyindole are not available,ionization potentials for these molecules were estimatedby a method based upon Koopmans’ theorem j37]. A linear regression between the experimental vertical ionization potentials (IP) of indole [ 361, benzene [ 381, naphthalene [ 381 and anthracene [ 381 and the energies of their highest occupied molecular orbital (&‘norwo),calculatedwith an LP3G basis set [ 291 (correlation coefficient, e0.998; significance level,
TABLE 1 Correl&ion between vertical ionization potential (IP) and energy of highest occupied molecular orbital (HOMO) Molecule
-
3enzene Na~hth~ene Antbraeene Indole
10‘20 6.99 8.02 8.70
9.24b 8.15b 7.4ob 7.90”
8.61 8.35 8.47
7.8Sd 7.64d 7.74d
5-HIND 6-HIND 5-HT
G~OMO~
WI
IP (eV)
“From SCF calcuiation with LP-3G basis set. bFromref. 38. “Fromref. 36. ‘~xtra~~~~orn thelinearregression: IP=0.524+0.852(-;ENoMo),
fr=0.998).
P
The molar enthalpy of subl~ation A.&z of a molecularcrystal at a temperature T is givenby f4 f.]
327
&SUB=EGAS-EPACK-&AT+PV
(10)
whereEGAs is the translationaland rotational energyin the gas phase (3RT), EpAcK is the crystal packing energy (lattice potential energy), ELAT is the kinetic energy associated with the lattice vibrations, and the PV term is taken to be equal to RT (neglecting the volume of the solid relative to that of the gas). The molecules are assumed to be rigid bodies; the internal vibrational energiesare assumedto be the same in both gas and solid phases. The crystal packing energyE PACKwas calculatedfrom the relation where EsCF(xl) is the SCF contribution to the lattice potential energy and EDIsp(xl) is the dispersioncontributionto the latticeenergy.In principle,EpACK should be determined by summing over all pairwise intermolecularinteractions in the crystal lattice. Since the magnitudeof SCF interactions is small for neutral non-polar molecules, SCF peculations, with BSSE correction, for benzene and naphth~ene were done only for one moleculeand its near neighbors. Dispersion energieswere calculated according to the Claverie [ 34 ] formulation using either Scheme I or Scheme II (see above). The lattice summation was done by choosing a reference molecule and calculating pairwise interactions with increasingly distant neighbors until the magnitude of the interactionbetween a pair of moleculeswas less than 0.02 kcal mol-‘. The kinetic energyof the lattice vibrationsELAT was calculatedfrom experimental and/or calculated lattice frequencies. The kinetic energy of optical modes was calculated according to the Einstein app~ximation [41]. The kinetic energy of acoustical modes was calculated according to the Debye approximation [41]. The lowest optical frequency was taken as the maximum frequencyvMAXin the Debye formulation. For a crystal whose geometryis known at a given temperaturefrom crystallographic data, determination of E PACKand EMT allows the calculation of A.&un from eqn. ( 10). The calculatedvalue of AHsunmay be compared with the experimentalenthalpyof sublimation.By usingeitherSchemeI or Scheme II in the dispersion energy contribution to E PACK,it is possible to determine which scheme is preferable (see below). Ilet&
of the electrostatic and dispersion seems
One molecule (5-HIND, 5-HT or 6-HIND) was kept fixed while the scanning molecule (indole) was oriented with its molecularplane parallel to that of the fixed molecule at an interplanardistance of 6.238 bohr. The indole portion of the fmed molecule was taken to be the xy plane with the origin at atom Nl of the indole ring. The y axis was directed along the Nl-C2 bond. The coordinate system is depicted in Fig. 1. The scanning moleculewas translated
q -7.0 0 -5.0 A -3.0
7
A I
I
-10.0
-5.0
5.0
1
I
I
0.0
10.0
15.0
X (bohr) a
s 0
ui 0
E
d
0
e >
0
Y 0
s
A
0 .
0
? I
-10.0
-5.0
I
0.0
I
I
5.0
10.0
1
15.0
X (bohr)
Fig. 1. Total interaction energy ICecaNbetween a fixed molecule (shaded) and a scanning molecule is calculated as the sum of electrostatic and dispersion energies, and is represented as isoenergy contours (kcal mol-I). The scanning molecule is in a parallel plane 6.238 bohr from the fixed molecule. Sketches of the scanning molecule are shown at positions corresponding to local energy minima. A. 5-HIND (fixed) and indole. B. 6-HIND (fixed) and indole.
along a square grid with a step size equal to 0.5 bohr. At each point, the indole was rotated counterclockwise in increments of 10” about an axis perpendicular to the molecular plane through the midpoint of the C&C9 bond (the position where the pyrrole and benzene rings are fused). At each point, values of Eas, EDIsp, the lowest interaction energy and the corresponding orientation angle were saved. Once the lowest energy had been determined for each point on the
329
grid,positions of local energyminima were found by comparison of the energy at a givenpoint with the energyat neighboringpoints. The interactionenergy for the entire scan was representedby means of isoenergycontour maps. The molecules5-HIND, 6-HIND and indole each have symmetryC,. Thus, scans of indole with 5-HIND or 6-HIND can have two distinct relativeorientations. In the “P” scan, the initial o~en~tion of the scanning indole superimposes it with the indole portion of either B-HIND or 6-HIND so that the angle between the respective C&C9 bonds is zero (scan angle, 0” ). In this orientation,the calculateddirection of the dipole vector of the scanningindole is 195’ with respectto the x axis. In the “M” scan, the initialindole orientation (scan angle, 0” ) was generatedby reflecting the starting orientation for the “P” scan through the yz plane which passes through the Nl-C2 bond. As a resultof this reflection,the correspondingdirection of the indole dipole is 345’ with respect to the x axis (see Fig. 1) . Since the side-chain of 5-HT is out of plane, the molecularsymmetryis C,. Thus, there are four possible scans of indole relativeto 5-HT. To avoid close contact between atoms of the two molecules,the only scans actuallyrun were the “P” and “M” scans with the indole on the side of the plane distal to the side-chain. RESULTS AND DISCUSSION
Comparison between calculated and e~~ri~ntal
AHsUB
The comparison between calculated and experimentalenthalpies of sublimation for benzene and naphthalenecrystalsis shown in Table 2. For benzene, the value of AHsus has been measuredat 273 K [ 421, but high resolutionneutron diffraction data [ 241, used to calculateAHsUB,have been measuredat 138 K and 218 K. Thus, calculated values of AHsoa at the last two temperatures were linearly extrapolated to 273 K so that comparison could be made with experiment. A similar extrapolative method was employed by Banerjee and Salem [43]. In the calculation of EMT (see the Meth~ section), fr~encies of lattice vibrationsfor benzene at 138 K [ 42 ] and naphth~ene at 298 K 144] wereused, Since experimentallattice frequencieswere not known for benzene at 218 K, the value of EUT was estimated from a linear regressionof ELATversus temperature.For several molecular crystals, previously reported lattice frequencies were used to calculate values of ELAT.The crystals used, along with the correspondingtemperatures,were benzene at 138 K [ 421, naphthaleneat 298 K [44 1, cu-p-C,H&l, at 300 K [41], /3-p-CGH4Clz at 318 K [41], pyraxine at 180 K and 300 K [42], a-s-triazine at 300 K [42], j&&iaxine at 150 K [42] and s-tetraxineat 253 K [ 421. The crystalslisted are composed of neutralnon-
polar aromatic’molecules of similar size. The equation for the linear regression of ELAT versus temperature (TEMP ) is E LAT=co +c,(TEMP)
(12)
A fit of the nine data points to this equation yields the coefficients c,=O.154 kcal mol-’ and c,=O.O115 kcal mol-’ K-l. The points are closely correlated (r= 0.999; P< 0.~1) - From this regression, the predicted value of EMT for a crystal at 218 K is 2.7 kcal molW1. As expected for neutral non-polar molecules, the magnitudes of EsCF(xl ) for benzene and naphthalene shown in Table 2 are small in comparison with the corresponding magnitudes of EnIsp (xl). Thus, EPAcx depends primarily on dispersion. Neutron diffraction structures for benzene [ 241 at 138 K and 218 K and X-ray diffraction structures for naphthalene [ 251 at room temperature (taken to be 298 K) were used in the calculation of dispersion energies. The TABLE 2
Comparison between calculated and experimental values of enthalpy of sublimation for benzene and napbthalene’ Molecule
138 1.1 1.7 0.4
T WI 4RT=EGAsSPV E b EZ(xl)C
218 1.7 2.7 -0.4
273
296 2.4 3.6 1.1
Schemed
EnIsP(xW
E PACKf W”B (talc 1s A&&XptP
I
II
I
II
- 16.4 - 16.0 15.4
- 13.4 - 13.0 12.4
- 14.9 - 15.3 14.3
-12.2 - 12.6 11.6
II
I -
-
13.5 10.7
11.1
I
II
-23.5 -22.4 21.2
- 19.0 -17.9 16.7 17.3
*Energies in kcal mol-‘. bKinetic energy of lattice cakulated from frequencies of lattice vibrations as described in text. Frequencies for benzene at 138 K from ref. 42. Erequencies for naphthalene from ref. 44. The vahre of .8&T for benzene at 218 K wss estimated from a linear regression (eqn. (12)). “SCF contribution to lattice potential energy corrected for BSSE (ref. 33). dScheme I: cyTfor C-CAROM= 0.59 A3. Scheme II: aT for C-Cmo,=0.40 A3 (see text). “Dispersion contribution to lattice potential energy calculated using eqn. (5). Total lattice potential energy calculated using eqn. (11). pEnthalpy of sublimation calculated using eqn. (10). Values for benzene at 273 K were obtained by liuear extrapolation from corresponding values at 138 K and 218 K. hExperimental enthalpy of sublimation. Value for benzene from ref. 42; value for naphthalene from ref. 45.
331
bond polarizabilitiesfor C-H bonds were taken as cyL=arT=0.64 A”. For aromatic C-C bonds, (Yewas taken as 2.21 A3and aT was taken as either 0.59 A” (Scheme I) or 0.40 A3(Scheme II). The mean excitation energy U for each moleculewas set equal to 1.8 times the experimentalverticalionization potential (see Table 1) . Thus, UBmz= 16.63 eV and UN-n = 14.67 eV. In each of the cases treated,Emsp(xl) makes the predominantcontributionto EPAcx.In fact, B&-&xl) could have been omitted from the collations ~thout introducing significant errors into the comparison between the calculated and experimentalvaluesof AI&n. It maybe seenin Table 2 that valuesof Msua calculatedaccordingto Scheme II ( aT for C-CAROM=0.40 A3)arein better agreementwith experiment for both benzene [ 421 and naphthalene [ 451. Consequently,Scheme II was used in all of the dispersion energycalculationsreportedbelow. ~~c~ost~t~ and dispersion stuns
Isoenergycontour maps of EscANfor the interactionof indole with 5-HIND (“P” scan), and 6-HIND (“P” scan) are presentedin Fig. 1. Parametersused TABN
3
Parameters used in the calculation of the dispersion energy Molecule
Mean excitation energy“ (kcal mol-‘)
Benzene Napht~ene Indole B-HIND 6-HIND 5-HT
383.64b 3383Ob 327.92” 326.2Sd 317.13d 321.28d
Bond
Bond polarixabilit~ (A3) Longitudii
C-H c-c C-GEi,,’
C-N c-o N-H O-H
0.64 0.99 2.24 0.57 0.89 0.50 0.58
‘1.8 times vertical ionization potential. bVertical ionization potential from ref. 38. cVertical ionization potential from ref. 36. dvertical ionization potential from linear regression (see Table 1) . “VaIues for O-H bond from ref. 40. Other values from ref. 39, ‘All bonds between heavy atoms in indole ring taken as C-C (arom). @I’rausverse~~~b~ity taken as mean of in-plene and out-of-phme vahres.
Transverse 0.64 0.27 0.408 0.69 0.46 0.83 0.79
332 TABLE 4 Positions, energies and orientations of local energy minima derived from electrostatic and dispersion scans with indole” “Fixed” molecule Scan type Position of minimumb Eae 5-HIND
“P”
“M”
B-HIND
“P”
5-HT
=P*,
( - 1.5, -3.5) ( 6.5, 6.5) ( 7.0, -7.0) ( -3.5, 0.0) (-1.5, -4.0) ( 5.5, 6.5) ( 6.5, -7.0) ( -2.0, -3.0) ( 11.0, -0.5) ( - 1.5, -3.5) ( 7.0, -7.0) ( 7.0, 6.5)
-3.56 -2.20 -3.72 -3.74 -3.59 -2.69 -3.47 -3.94 -3.69 -3.72 -3.69 -2.87
EDrsp EsCAN Dipole angle” (deg) -4.43 -3.62 -3.67 -3.79 -4.07 -3.70 -3.81 -4.23 -3.79 -4.48 -3.67 -4.72
-7.99 -5.82 -7.39 -7.53 -7.66 -6.39 -7.28 -8.17 -7.48 -8.20 -7.36 -7.59
25 265 95 25 55 265 115 15 175 25 95 255
“Positions in bohr, energies in kcal mol-‘, orientations in degrees. bx-y coordinates of midpoint of C&C9 bond in scanning indole. ““P”scan: (dipole angle) = 195” + (scan angle); “M”ecan: (dipoleangle) = 345” + (scan angle).
to calculateEDIsPfor these and other scans are given in Table 3. A sketch of the fixed molecule is shown in each map along with a sketch showingthe position and’orientation of the indole at each local energy minimum. Table 4 gives the position and orientation of each local energy minimum along with the values of EscAN,Eoo and EnIsp.In addition to data for the scans shown in Fig. 1, Table 4 also contains data for two other scans (5 HIND “M” and 5-HT “P”). In comparing the two scans involving &HIND, one observes that for each local rni~~urn in the “P” scan there is a correspondingminims in the “M” scan with a similarposition and energy.In addition,the dipoles of a pair of correspondingminima (one “P”, one “M”) have similarorientations.Similar trends are observed when comparingthe “P” and “M” scans for 5HT or 6-HIND (data not shown). The position, orientationand energyof each local minimumin the 5-HIND scan closely correspond to a minimum in the 5-HT scan. This finding thus supportsthe use of 5-HIND as a model for 5HT, especiallysince the scan of 5-HT with indole was done on the side of the XYplane away from the 5-HT side-chain. The “P” scan with 5-HIND is quite different from that with 6-HIND (see Fig. 1) . Only the minimanear the pyrroleportion of the furedmoleculesare in close agreement.Each map is characterizedby a region of weak interaction which spans the indole ring and the hydroxyl group. The difference in the positions of the O-H group leads to a large difference in the characterof the contour maps. When the scanning molecule is close to the oxygen atom, the
333
magnitude of EscAN is small. As a result of the pronounced effect of the oxygen atom on the shape of the contour map, the position of the local minimum at (7.0, - 7.0) in the &HIND scan corresponds to a position of very weak interaction in 6-HIND. For this minimum in the &HIND scan, the value of .&CAN is - 7.39 keal mol- l at an orientation angle of 260’. For the same position in the 6-HIND scan, the value of EscAN is -1.61 kcal mol-’ at a similar orientation angle of 290”. This difference of 5.78 kcal mol-” in the interaction energy comes p~m~ily from the electrostatic infraction. For 5-HIND, EQa is -3.72 kcal mol-’ and Emsp is -3.67 kcal mol-l. The corresponding values for the 6-HIND are + 2.45 kcal mol-* and -4.06 kcal mol-I, respectively. Thus, the difference in EQa is responsible for the selectivity demonstrated by the model of a neutral recognition site in the receptor. If one assumes that the geometry of B-HIND at the local minimum identified by the scan corresponds to a high-affmity binding configuration of 5HT at a 5-HT receptor, then 6HT would be expected to have a much lower affinity for that receptor because it will have an ~favor~le electros~tic interaction with the receptor site. In fact, experimental measurements show a large difference in affinities between 5-HT and 6-HT at both 5-HTlA and 5-HT2 receptors. Binding studies at 5 HTXAreceptors [46] give dissociation constants of 3.1 nM for 5HT and 1600 nM for 6-HT. According to measurements of 5HTz receptors in the isolated rabbit aorta [47], the dissociation constant for 5HT is 250 nM, whereas the dissociation constant for 6-NT is greater than 10 000 nM,
Since the scanning position at (7.0, - 7.0 ) clearly distinguishes between 5-HIND and 6-HIND, the configurat,ion from the “P” scan of 5HIND with indole at that position was selected for a de~led investigation of the variation in scan energies with the orientation angle. The results are shown in Fig. 2. The magnitude of E nIsP is generally greater than that of I&o, and the variations in Eoo and Enrsp are almost exactly out of phase. The most favorable total infraction energy occurs at an orien~tion angle of 260” between the C& C9 bonds of the two molecules. In this orientation, both the electrostatic and dispersion terms are attractive and contribute equally to the stabilization of the complex. Since the two molecules do not overlap appr~iably (see Fig. 1A ) , the electrostatic interaction is at a minimum and the dispersion interaction is at a maximum. On the other hand, maximum overlap of the two rings (which occurs at an orientation angle of 150” between the (X-C9 bonds) gives rise to a repulsive electrostatic energy and an attra~ive dispersion energy (see Fig, 2 ). The angular variation in the total interaction energy, EscA, follows that of IZoo because Eee varies over a range of approximately 4.8 kcal mol-‘, while EnIsp varies only over a range of approxima~ly 1.5kcal mol-“. Consequently,
334
0
ELEC
A DISP
404 0
I
I
I
I
I
60
120
180
240
300
ORIENTATION
ANGLE
I 360
(degrees)
Fig. 2. Dependence of interaction energies between &HIND and indole on the orientation angle between the C&C9 bonds of the two molecules. The relative position of the two molecules is at local minimum (7.0, -7.0) as shown in Fig. IA. ELEC, electrostatic energy; DISP, dispersion energy; TOTAL, total interaction energy.
the electrostaticinteractionis the importantproperty in determiningthe preferred orientation and in establishing a molecular correlate of selectivity, whereasthe dispersion energyprovides the major portion of the stabilization of the complex. Hunter and Sanders [48] reached a similar conclusion in a recent analysisof 7c-xinteractionsin porphyrins. Similar orientations properties were found in scans in which i~~olium was used as a model of a receptor that contains a chargedresiduein it [f&10]. However, the mechanismsfor selectivity and affinity seem to be different in the two models. In the chargedmodel both the interactionenergy,a correlate of affinity, and the orientation of the interactingmolecules, a basis for selectivity, were determinedby electrostatics. In the neutral model, which is explored in this work, the electrostaticinteraction governsthe orientationalselectivitywhereasthe dispersioninteractioncontributesmainly to the affinity. Furthermore,in the charged model the orientational selectivity was determined by the favorable interaction between the ~stribution of the positive charge in irni~o~~ and the electros~tic potential of 5HT, whereas the interactionbetween the neutralindole and 5-HT may be better characterized by the interactionbetween the indole dipole and the electric field of 5-HT. The interactionenergyEWbetweenthe indole dipole p and the 5-HT electric field Eis givenby Ep=
-~-ST
(13)
Because the electric field Eis the negative gradient of the molecularelectrostatic potential of 5-HT, Fig. 3 showsthe electrostaticpotential of 5-HT in the
335
IJ -16.0 O -12.0 A -8.00 + -4.00
0
+ 0
:
s 0 -10.0
-5.0
0.0
5.0
10.0
15.0
f;; *
20.0
X (bohr)
Fig. 3. Dipole vectors of indole at local minima obtained from a scan of 5-HT and indole are superimposed on the molecular electrostatic potential map of 5-HT calculated in a plane 6.238 bohr from the plane of the indole portion (see the Methods section). The direction of each dipole vector is approximately aligned with the electric field generated by 5-HT, which is perpendicular to the equipotential contours (kcal mol-’ ) .
plane (z= -6.238 bohr ) in which the electrostaticand dispersion scans were carried out. Sketches of the local minima for indole obtained from the 5HT “P” scan are superimposedon the electrostaticpotential map. For each indole, the dipole vector is shown at the midpoint of the bond between C8 and C9. It may be seen that the direction of each dipole vector is approximatelythe same as the direction of the electric field of 5-HT, which is perpendicularto the contours. At each local minimum,the indole molecule is located in a region in which the electric field is relativelystrong and uniform, i.e. a region where the equipotential contours are closely spaced and approximatelystraight. Thus, the dipole-electric field interaction model may be used to interpret the electrostatic and dispersionscans in Fig. 1. Stronginteractionsoccur in regionswhere 5-HIND or 6-HIND has a strongelectricfield. At (7.0, - 7.0), a position where the interaction energywith indole clearly distinguishesbetween &HIND and 6-HIND, the electricfield of 5-HIND is strongbut the field of 6-HIND is weak. Interaction energies of 5-HIND complexes
The calculated interaction energiesfor the complex of B-HIND and indole at the (7.0, -7.0) position from the “P? scan are presented in Table 5. The interplanardistance is 6.238 bohr.
336 TABLE 5 Camparison of interaction energies’ in &HIND complexesb B-HIND and indole “P” (7.0, -7.0)
&HIND and imidazolium (7.5,3.0)
-3.72 -3.67
-11.97
- 7.39
&C.&N=&QfhSF’
SCE Morokuma decomposition -11.86 1.62 -0.94
ES EX
- 3.90 1.22
PL CT MIX
-0.35 -0.23 0.02
=%m
- 3.24
- 11.61
&xxfco~)
-2.70
-11.11
&wr~t”&w
(corr) +EDISP
-0.46 0.03
-6.31
*Energies in kcal mol-‘. ‘~~~~ distance is 6.238 hohr, cEm~ could not be c&&ted for ~i~~li~ ities af charged molec&~ are not available.
complex because parameters for bond polarizabil-
accordingto the scheme The SCF ~bi~tion energy,EscFwas diurnal of Morokuma [30,31] . The value of E s& was corrected for basis set superposition error [ 331 to givea quantitydesignatedas EeCF(corr). Interestingly,the valueof je,, is in close agreementwiththe valueof the electrostaticterm (ES) a Thus, at this interplanardistance, the monopole-monopole approximationof the electrostaticinteraction gives a reasonable representationof the electrostatic energy. The EB1sPaccounts for 58% of EmYAL,the total s~ili~ation energy.Thus, tbe s~b~ity of the complex between &HIND and indole has a ~onsiderahle~ontributioufrom the dispersion in~~~~ion. It is of interest to compare the Moro~ma decomposition of the energy of i~~ra~tiou in the complex of G-HINDand indole with that in the complex of &HIND and imidazolium cation (see Table 5). Indole represents a neutral model for a 5-HT receptor,whereasimidazoliumrepresentsa chargedmodel. The local minimumconfigurationsof the 5-HIND-imidazolium complex were determinedfrom a mo~opole-mo~opole scan, using LP-3G charges,at an interplanardistance of 6.236 bohr. The local minimumat position (?.S, 3.0) in the coordinate system used for the ~-HIP-~dole scans correspmds to that
337
obtained previously in similar scans of B-HT with imidazolium [ 101. Similar to the alignment of the dipole vector of the scanning indole with the electric field of 5-HT (see Fig. 3), we find that the direction of the dipole vector of imidazolium also coincides with the direction of the electric field of &HIND. The most important difference between the two complexes is in the SCF interaction energies. In the complex with imidazolium, EsCF is - 11.61 kcal mol-’ whereas in the complex with indole it is only -3.24 kcal mol-‘. The decomposition of the interaction energy in the 6-HIND-imidazolium complex provides an explanation for this large difference. The magnitude of ES in the 5HIND-imi~zolium complex ( - 11.86 kcal mol-‘1 is much larger than the correspon~ng value in the 5-HID-indole complex ( - 3.90 kcal mol-l ). The ma~i~des of EX (1.62 kcal mol-” 1, PL ( - 0.94 kcal mol-l ) and CT ( -0.46 kcal mol-’ ) are similar to the correspondingvalues for the indole complex (see Table 5). We also note that although the decomposition of EsCF for the two complexes is quite different, the BSSE is approximately 0.5 kcal mol- ’ for each complex. Thus, it appears that the BSSE reflects the size of the molecular basis set rather than the nature of the infraction between &HIND and the respective receptor models. In the imidazolium complex, the contribution of the electrostatic term to the SCF energy is the main stabilizing factor, whereas in the indole complex dispersion plays an important role, contributing over 50% to the interaction energy. The methods used here to calculate dispersion energies could not be applied to the imidazolium complex because parameters for bond polarizabilities of charged molecules are not available. However, the polar~bi~ty of the imidazolium cation is expected to be small, and ~onse~ently the dispersion interaction energy of the imidazolium complex is expected to be less than that of the indole complex.
The changes in electron density upon formation of the complex between 5HIND and indole at the (7.0, - 7.0) position from the “P” scan were studied in two related ways. Values of Ap for the complex with Az equal to 6.238 bohr, corrected for BSSE and calculated by integration in planes parallel to the molecule planes (see the Methods section), are plotted in Fig. 4. The figure shows a depletion of electron density midway between the indole planes. This probably arises from repulsion between the electron clouds of S-HIND and indole. Simul~eously, the electrons in each molecule are polarized in a direction perpendicular to the indole plane, so that the negative end of each induced dipole points toward the region between the planes. In this manner, the electron charge of each molecule of the complex is polarized towards the other molecule. It may be seen in Fig. 4 that the polarization of 5-HIND is slightly greater than that of indole. The
INDOLE
4.23*
DISTANCE
S-HIND
0.000
(bohr)
Fig. 4. Electron density (e/bohr X 103) redistribution in the (B-HIND + indole) complex at local minimum (7.0, - 7.0) (see Fig. 1A) calculated from molecular wavefunctions obtained with the LP-3G basis set. 4p is calculated by integration in planes parallel to the molecules in the complex and~~~ed~~(~mplex)-~(5-H~+gh~t ~dole)-~(indole+gh~t 5-HIND). See the Methods section for details.
overall pattern of the charge redistributionexplains why dispersion,which is a correlated interaction between induced dipoles, makes a large contribution to the total stabilizationenergyof the complex (see Table 5). The electron density redistributionin the complex of S-HIND and indole contrasts sharply with that in the complex of 5-HT and imidazoliumcation which was reported previously [ 81. In the latter case, 5-HT was more extensively polarized (toward the imi~oli~) and the imi~oli~ was slightly polarized away from the 5-HT. In the ~i~zoli~ complex, 5HT was polarized nearly eight times more than 5-HIND in the indole complex. The depletion of electron density midway between the molecule planes was relatively small.The polarizationpattern in the 5-HT-imidazolium complex was shown to be characteristic of a strong electrostatic interaction. The difference between the polarization in the imidazolium and indole complexes is also reflected in the difference between the PL terms in the corresponding energy decompositions (see Table 5). A more detailedanalysisof the electron density redistributionwas obtained by computingdifference maps in planes parallelto the indole planes. Figure5 shows maps for the planes at a distance of 1.9 bohr on either side of each molecule. Thus, Figs. 5A and 5D represent charge redistributionsin planes outside the molecularcomplex and Figs. 5B and 5C) in planes inside the molecular complex. Each of the selected planes corresponds approximatelyto a a-electron region in one of the molecules.In all four planes, there is a distinct polarization of electron density away from the region in which the molecules overlap.This polarizationis moreextensivein the spacebetweenthe molecules as can be seen from a comparison of Figs. 5B and 5C with Figs. 5A and 5D.
339
0
0
vi-
vi
0
0
c d
d
‘>
9
T
0
80
*
?_
Lp
‘f 0
s-
q
0
-0.30
_
0
-0.30
s D s
+ -10.0
-5.0
0
A
0.30 I
I
0.0
I
5.0
I
10.0
I
?
15.0
X (bohr)
B -10.0
I
-5.0
I
0.0
I
5.0
I
10.0
/
15.0
X (bohr) d
X (bohr)
X (bohr)
Fig. 5. Electron density (e/bow X 104) redistribution in the (5-HIND + indole) complex at local minimum (7.0, - 7.0) (see Fig. 1A) calculated from molecular wavefunctions obtained with the LP-3G basis set in planes parallel to the indole portions of the molecules. (B-HIND in plane z = 0; indole in plane z = - 6.238 bohr ). Shaded regions correspond to increases, and unshaded regions to decreases in electron density upon complex formation. Each selected plane corresponds approximately to a x-electron region in one of the molecules. A. z = + 1.900 bohr. B. z = - 1.900 bohr. C. z= -4.338 bohr. D. z= -8.138 bohr.
This charge redistribution reduces the repulsion between the electron clouds of the two molecules, and thus stabilizes the complex by inducing oppositely directed dipoles parallel to the planes of the two molecules (compare Fig. 5A with Fig. 5D and Fig. 5B with Fig. 5C). The formation of these induced dipoles gives additional support for the importance of polarizability in the interaction complex between 5-HIND and indole.
CONCLUSIONS
We haveexplored the possibility that the recognitionof 5-HT by two different receptors is governed by different physical interactions. It was demonstratedthat 58% of the interactionenergyof 5-HT with a neutralligand,modeled by the indole portion of tryptophan, was due to dispersion;the rest was mostly due to elect~static infractions. This is in clear contrast to the previously studiedinteraction of 5-HT with imidazoliumcation [ 8-10 1, where the’ interaction was found to be primarily electrostatic. In spite of the different interactionsthat contributeto the stabilityof the complexes,the most important element in the recognition of 5-HT by both of these receptor models is electrostatic.With indole servingas the model, the recognition is based on its polarity, and is expressedin the interactionof its dipole with the electric field of &NT. With imidazolium,on the other hand, the recognitiondepends upon the interactionbetween the distributionof its positive chargeand the electrostatic potential of 5-HT. Assessmentof the roles that these models (indole and imidazo~um) play in 5-HT receptorsrequiresadditionalexperimentalinformationabout the structures of the receptorproteins. In the meantime,we can determinewhetherthe demonstratedselectivityof differentdrugstowardsthe known 5-HT receptors can be correlatedwith the distinguishingproperties of the models. The different mechanismsthat are responsible for affinity-dispersion in the neutral model and electrostatic in the charged model-illustrate a possible basis for the specificityof ligandstowards the various 5-HT receptors. ACKNOWLEDGMENT
We thank Dr. Hare1Weinsteinfor his helpful comments on the manuscript. Computations were performed on the supercomputersystem at the Cornell National SupercomputerFacilitywhich is sponsored by the National Science Foundation.The authorsgratefullyacknowledgethis resourceand the generous allocationsof computertime from the UniversityComputingCenterof the City Universityof New York. This work was supported by the National Institute on Drug Abuse under grant DA-01875 and by the National Institute of General Medical Sciences under grant GM-34852. REFERENCES 1 2 3
J.P. Green and S. Maayani, in J.W. Black, D.H. Jenkinson and V.P. Gerskowitch (Eds. ), Perspectives on Receptor Classification, Alan R. Liss, New York, 1987, pp. 237-267. S.J. Peroutka, Annu. Rev. Neurosci., 11 (1988) 45. A. Shenker, S. Maayani, H. Weinstein and J.P. Green, Mol. Pharmacol., 31 (1987) 357,
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M. DeVivo and S. Maayani, J. Pharmacol. Exp. Ther., 238 (1986) 248. B.L. Roth, T. Nakaki, D.-M. Chuang and E. Costa, Neuropharmacology, 23 (1984) 1223. D. Hoyer, G. Engel and H.O. Kalkman, Eur. J. Pharmacol., 118 (1985) 13. H. Weinstein, D. Chou, S. Kang, CL. Johnson and J.P. Green, Int. J. Quantum Chem., Quantum Biol. Symp., 3 (1976) 135. H. Weinstein, R. Osman, W.D. Edwards and J.P. Green, Int. J. Quantum Chem., Quantum Biol. Symp., 5 (1978) 449. H. Weinstein, R. Osman, S. Topiol and J.P. Green, Ann. N.Y. Acad. Sci., 367 (1981) 434. H. We~s~in, R. Osman, J.P. Green and S. Topiol, in P. Politzer and D.G. Truhlar (Eds. ), Chemical Applications of Atomic and Molecular Electrostatic Potentials, Plenum, New York, 1981, pp. 309-334. R. Osman, H. Weinstein, S. Topiol and L. Rubenstein, Clin. Physiol. Biochem., 3 (1985) 80. R. Osman, S. Topiol, L. Rubenstein and H. Weinstein, Mol. Pharmacol., 32 (1987) 699. G.A. Mercier, R. Osman and H. Weinstein, Protein Eng., 2 (1988) 261. G.A. Mercier, R. Osman and H. Weinstein, in R. Rein and A. Golombek (Eds.), ComputerAssisted Modeling of Receptor-Ligand Interactions: Theoretical Aspects and Applications to Drug Design, Alan R. Liss, New York, 1989, pp. 399-410. D.D. Jones, J. Theor. Biol., 50 (1975) 167. T.T. Takigawa, T. Ashida, Y. Sasada and M. Kakudo, Bull. Chem. Sot. Jpn., 39 (1966) 2369. A. Wakahara, M. Kido, T. Fujiwara and K. Tomita, Bull. Chem. $0~. Jpn., 46 (1973) 2475. R.S. Root-Bernstein and F.C. Westall, Brain Res. Bull., 12 (1984) 425. P.R. Carnegie, Nature, 229 (1971) 25. E.H. Eylar, S. Brostoff, G. Hashim, J. Caccam and P. Burnett, J. Biol. Chem., 246 (1971) 5770. G. Falkenberg, Thesis, Karolinska Institutet, Stockholm, 1972. D. Carlstrom, R. Bergin and G. Falkenberg, Q. Rev. Biophys., 6 (1973) 257. E.G. Cox, D.W.J. Cruickshank and J.A.S. Smith, Proc, R. Sot. London, Ser. A, 247 (1958) Z.E. Bacon, N.A. Curry and S.A. Wilson, Proc. R. Sot, London, Ser. A, 279 (lQ64) 98. D.W.J. C~icks~k, Acta Crystallogr., 10 (1957) 504. D.W.J. Cruickshank, Acta Crystallogr., 9 (1956) 915. J.S. Binkley, R.A. Whiteside, R. Krishnan, R. Seeger, D.J. DeFrees, H.B. Schlegel, S. Topiol, L.R. Kahn and J.A. Pople, GAUSSIAN 80 IBM version, unpublished. S. Topiol, J.W. Moskowitz and CF. Melius, J. Chem. Phys., 70 (1979) 3008. S. Topiol and R. Osman, J. Chem. Phys., 73 (1980) 5191. K. Morokuma, J. Chem. Phys., 55 (1971) 1236. K. Kitaura and K. Morokuma, Int. J. Quantum Chem., 10 (1976) 325. S. Srebrenik, H. Weinstein and R. Pauncz, Chem. Phys. Lett., 20 (1973) 419. S.F. Boys and F. Bernardi, Mol. Phys., 19 (1970) 553. P. Claverie, in B. Pullman (Ed.), In~~o~~ar Interactions: From Diatom& to Biopolymers, Wiley, New York, 1978, pp. 69-305. S. Scheiner and M.M. Szczesniak, Int. J. Quantum Chem., Quantum Biol. Symp., 11 (1984) 201. J.H.D. Eland, Int. J. Mass Spectrom. Ion Phys., 2 (1969) 471. ‘I’. Koopmans, Physica, 1 (1933) 104. P.A. Clark, F. Brogli and E. Heilbronner, Helv. Chim. Acta, 55 (1972) 1415. R.J.W. Le Fevre, Adv. Phys. Org. Chem., 3 (1965) 1. M.M. Szczesniak and S. Scheiner, J. Chem. Phys., 80 (1984) 1535. H. Bonadeo, E. D’Alessio, E. Halac and E. Burgos, J. Chem. Phys., 68 (1978) 4714. Z. Gamba and H. Bonadeo, J. Chem. Phys., 75 (1981) 5059. K. Banerjee and L. Salem, Mol. Phys., 11 (1966) 405.
44 45 46 47 48
S. Califano, R. Righini and S.H. Wahosley, Chem. Phys. I&t., 64 (1979) 491. R.S. Bradley and T.G. Cleaeby, J. Chem. Sot., (1953) 1690. G. Engel, M. Gothert, D. Hoyer, E. Schlicker end K. Hillenbrand, Naunyn Schmiedebergs Arch. Pbarmakol., 357 (1986) 1. B.M. Clancy and S. Maayani, J. Pharmacol. Exp. Ther., 233 (1985) 761. C.A. Hunter and J.K.M. Sanders, J. Am. Chem. Sot., 112 (1990) 5525.
321
Elsevier Science Publishers B.V., Amsterdam
The interaction between 5-hydroxytryptamine tryptophan: a serotonin receptor model
and
Lester A. Rubenstein”and Roman OsmarPb Departments of Physiology and Biophysics” and of Pharmucokgy’, Mount Sinai School of Medicine of the City Uniuersity of New York, New York, NY 10029 (USA)
(Received 5 November 1990)
Abstract A system consisting of serotonin f5-hydroxytryptamine, 5H’I’) and.tryptophan was used to study the process of recognition at a 5-HT receptor. The optimal geometries of interaction between 5-hy~x~ndole (5-HIND), a model ,of 5-HT, and indole, a model of tarpon, were obtained from a scan which included both eiectrostatic and dispersion energies. Recognition of 5-HIND by indole is determined by the electrostatic interaction between the components of the complex. Formation of such a complex results in electron density redistribution8 in directions both perpendicular and parallel to the planes of the interacting molecules. The nature of the charge polarization is consistent with the dominant contribution that dispersion makes to the stabilization energy of the complex. The optimal geometry of interaction between 6-HT and indole is different from the 5-HT geometry, both in the relative position of the com~nen~ of the compiex and in their orientation. This difference can be a basis for the selectivity of a receptor towards different liganda. Furthermore, the characteristica of the interaction of 5-HT with a charged imidaxolium cation, which were explored in our previous work, are different from those of the interaction of 5-HT with a neutral indole as a receptor model. In both cases, the electrostatic interaction is the o~en~tion~ determinant. With the imidaxolium cation, the main stabilization cornea from electrostatics. With the neutral indole, it comes from dispersion. Such distinctions ilhzetrate a possible basis for the specificity of ligands towards the various 5-HT receptors.
INTBODUCTION
Many ne~otr~s~t~r and drugmoleculescontain aromatic portions.Tbese portions play an important role in the selectiverecognitionof these molecules by complementaryresiduesin the receptor protein. The complementaritycan be describedin terms of the type of interaction betweenspecificportions of the receptor and the aromatic portion of the ligand. Quite often one neurotransmitter can be recognizedby different receptors. For example, serotonin (5hy~ox~~~ine, 5-HT ) is a neurotransmit~r whichacts at multiplereceptors [ 1,2], Its action on these receptors leads to different physiologicalresponses [ 3-51. The binding selectivity associated with the various receptors 0166-12~/91/$03.50
0 1991Elsevier Science Publishers B.V. All rights reserved.
[61 suggests that the mole~~&~ structures of the active sites may differ. Thus, a possible contribution to the selectivity of Merent receptors which recognize the same ligand (5-HT) may be the distinct physical nature of the interaction between ligand and receptor. In a model for a 5-HT receptor which has bsen described previously [ ‘7-10J, two elements of recognition of serotonin have been identified. The first element is an ionic interaction between the protonatedaminoethyl side-chain and a negatively charged site in the receptor. The second element is an interaction between the planar indole ring and a complement part of the receptor, which is represented by an imidazolium cation (the aromatic ring in the pro&mated form of histidine ) . Analysis of the stabilization energy of the complex between 5HT and imidazolium showed that the interaction is primarily electrostatic, and is enhanced by rn~~~ pola~~~on. Based on the changes in the charge ~tribution induced in the imidasolium cation upon its infraction with 5=HT, a proton transfer process from ~i~o~~ to a proton acceptor has been proposed f l&12 ] as a molecular m~hanism for the activation of a 5-HT receptor. The effect of a protein environment on this process has been studied in a&inidin, a protein of known structure with an appropria~ jux~~sition of the groups that form the proton transfer system proposed as an activation mechanism [ 13;14]. The indole ring in 5-HT may also interact with a neutral aromatic residue in the receptor. The electrostatic interactions between 5-HT and a neutral molecule are expected to be weaker than those with irni~~li~ cation but dispersion forces should also make an appreciable contribution to the interaction energy. The highly pol~i~ble indole ring in 5HT should give rise to favorable dispersion infractions with the pola~ble aro~tie amino acids phenyl~~ine~ tyrosine and trophy. Trophy, the most polarizabie and most polar of these three amino acids f15ftshould not only give an app~~iable dispersion interaction but should also functian as a sensitive electrostatic probe in its interactions with 5-HT. Accordingly, we have chosen to study the characteristics of the interaction between tryptophan and S-MT. Our choice of tryptophan was also guided by several experimental observations, First, close packing of indole rings is known to occur in crystals of tryptophan [X6] and 5hydroxytryptophan [ 171. Second, nuclear magnetic resonance studies of the binding of serotonin to severai oligopeptides indicate that t~toph~ undergoes a favorable infraction with 5-HT 1.IS].The bin~ng site in the peptide, co~spon~ng to residues 111-121in myelin basic protein [ 19,20], contains the essential sequence (Phe Ser Trp). A similar site in iuteinizing hormone-releasing hormone contains the sequence (Tyr Ser Trp). The binding sites in me~otropi~-stimulating hormone and adrenocorticotropic hormone each contain the sequence (Phe Arg Trp ). Thus, each of these 5HT binding sites consists of two aromatic residues fTrp, and either Phe or Tyr 1 separated by a hy~philic residue f&r or Arg ). The NMR data [ 18]
suggestthat the indole portion of 5-HT intercalatesbetween the two aromatic residues. In this study we have characterizedthe nature of the interaction between tryptophan and 5-HT and its dependence on the relative orientation of the molecules. We show that the interaction of 5-HT with a neutral molecule is different from its interactionwith a chargedcation and can serveas a basis for the selectivitybetween different 5-HT receptors. METHODS
Molecuhr geometries To reducethe size of the manta-mech~ic~ c~culations of the complexes between tryptaminesand trophy, the indole portions of these molecules wereused as models.The indoleportions of indob, 5hydroxyindole (5-HIND), 5-hydroxytryptamineand 6-hydroxyindole (6-HIND) were calculated in the Falkenberg averaged geometry [21,22]. Based upon previous findings [ 73, 5HT was calculated in the neutral form with the side-chain in the fully extended conformation. Geometriesfor benzene [ 23,241, naphthalene [ 251 and anthracene [26] were taken from crystallographicdata. Complexes between indole and !%HIND,indole and 5-HT, and indole and 6-HIND were studiedin configurationsin which the indole planes of each pair of moleculeswere kept parallel. Quantum-mechniccd
c&ulutins
Se~~cons~~nt field (SCF) c~cu~tions were done with the GAUSSI~ 80 systemof programs 1273. To reducethe numberof basis lotions, an effective core potential (ECP) [ 281 was used in conj~~ion with an LP-3G basis set [ 29 1, which was especially designed for the ECP. The interaction energy between the component molecules in a supermoleculewas decomposed by the method of Morokuma and co-workers [ 30,311 into contributionsfrom electrostatic interaction of the unperturbedmolecules (ES), mutualpolarization of the molecules (PL), exchangerepulsion(EX), chargetransfer (CT), and other residualenergycontributions (MIX). The molecularelectrostaticpotential of 5-HT was calculated from the molecular orbitals by a method described previously [ 321, and the resultswere representedas an equipotentialcontour map in a plane parallelto the indole. Electron density redistributions associated with complex formation were computed by subtracting the densities of the separated molecules from the density of the complex. The density differenceswere representedin two ways. In one method [8], density differenceswere computed in planes parallelto the indole portions by integratingthe electron density distributionover the plane.
324
The density differences were then presented as a plot of integrated density difference versus position of the plane. In a second method, density differences were calculated at points on a square grid in planes parallel to the molecules and displayed as isodensity contour maps. Calculations of stabilization energy and density difference were each corrected for basis set superposition errors (BSSE) according to the counterpoise method [ 33 1. The corrected stabilization energy, hEbB, and the corrected density difference, Ap-, are given by AE*n (corr ) = EAB-E(A+ghost)
-E(B+ghost)
Apm(corr) =PAB-P(A+ghost)
-p(B+ghost)
(I)
(2)
where AB represents the supermolecule, A and B the monomers, and a property of a given monomer is calculated using both its own basis set and the basis set of the other monomer (the ghost). Electrostatic and dispersion scan Supermolecule configurations (e.g. a complex of 5-HIND and indole) at local minima were obtained from a scan which included both electrostatic and dispersion interactions. The total interaction energy EsCANfor a given configuration of a dimer is given by E SCAN=E~~+E~I~~
(3)
where EQQdesignates the electrostatic energy, and EDIspthe dispersion energy. The electrostatic interaction energy was calculated with a monopole-monopole approximation using atomic Mulliken charges obtained from an SCF calculation with an LP-3G basis set [ 291. The electrostatic energy EQQis then given by EQQ=C
C i
j
QiQjlrG
(4)
where Qi and Qj are the Mulliken charges on the separate molecules A and B, respectively. The same sets of charges were used throughout each scan. rG is the distance between nucleus i in molecule A and nucleus j in molecule B. The dispersion energy was calculated from the interaction of bond polarizabilities according to the method of Claverie [ 341. The dispersion energy EDIsp is given by D’SP=4(UA+UB)
m ,,
(&J6
where, assuming the bonds to be axially symmetric
(5)
325
(6) VAand Unare the mean excitationenergiesof moleculesA and B, respectively. The subscriptm desi~a~s a bond in molec~a A, and the subsc~pt n a bond in molecule B. R,,= ]I$,, 1 where It,, denotes the vector joining the midpoints of bond m and bond n. A, and A, are the polar&abilitytensors of the two bonds, and T,, denotes the tensor TXn,=3(~,,X~,*)-~
(7)
where
(8) and aT is the transversecomponent of the bond polarixability.The anisotropy S of the pol~zability is given by &Z ffL-a;r
(9)
where aL is the longitudinalcomponent of the bond polarizabilityand emand e, are unit vectors in the directions of bonds m and n, respectively. Determination ofparameters for ~~u~t~~
of dispersion in~r~t~~
energy
A major cont~bution to the interaction energy between neutral molecules comes from the dispersionterm. The method of Claverie [ 341 chosen for these calculations requires the calibration of parameters for bond polarixabilities and molecular ionization potentials. Bond ~lari~bilities were calibrated by comp~ing calcula~d heats of sublimation for crystals of benzene and naphthalenewith experimentalvalues. Molecularionization potentialswere either taken from experimentor derivedfrom a linearregressionbetweenexperimental ionization potentials and calculated energiesof the highest occupied molecularorbital. For each molecule,the mean excitation energy, U, was taken to be 1.8 times the vertical ionizationpotential. This scale factor is the same as one used previously [ 351. The verticalionizationpotential of indole was taken from experimental data [SS]. Since experimentalionization potentials for 5- and 6-hydroxyindole are not available,ionization potentials for these molecules were estimatedby a method based upon Koopmans’ theorem j37]. A linear regression between the experimental vertical ionization potentials (IP) of indole [ 361, benzene [ 381, naphthalene [ 381 and anthracene [ 381 and the energies of their highest occupied molecular orbital (&‘norwo),calculatedwith an LP3G basis set [ 291 (correlation coefficient, e0.998; significance level,
TABLE 1 Correl&ion between vertical ionization potential (IP) and energy of highest occupied molecular orbital (HOMO) Molecule
-
3enzene Na~hth~ene Antbraeene Indole
10‘20 6.99 8.02 8.70
9.24b 8.15b 7.4ob 7.90”
8.61 8.35 8.47
7.8Sd 7.64d 7.74d
5-HIND 6-HIND 5-HT
G~OMO~
WI
IP (eV)
“From SCF calcuiation with LP-3G basis set. bFromref. 38. “Fromref. 36. ‘~xtra~~~~orn thelinearregression: IP=0.524+0.852(-;ENoMo),
fr=0.998).
P
The molar enthalpy of subl~ation A.&z of a molecularcrystal at a temperature T is givenby f4 f.]
327
&SUB=EGAS-EPACK-&AT+PV
(10)
whereEGAs is the translationaland rotational energyin the gas phase (3RT), EpAcK is the crystal packing energy (lattice potential energy), ELAT is the kinetic energy associated with the lattice vibrations, and the PV term is taken to be equal to RT (neglecting the volume of the solid relative to that of the gas). The molecules are assumed to be rigid bodies; the internal vibrational energiesare assumedto be the same in both gas and solid phases. The crystal packing energyE PACKwas calculatedfrom the relation where EsCF(xl) is the SCF contribution to the lattice potential energy and EDIsp(xl) is the dispersioncontributionto the latticeenergy.In principle,EpACK should be determined by summing over all pairwise intermolecularinteractions in the crystal lattice. Since the magnitudeof SCF interactions is small for neutral non-polar molecules, SCF peculations, with BSSE correction, for benzene and naphth~ene were done only for one moleculeand its near neighbors. Dispersion energieswere calculated according to the Claverie [ 34 ] formulation using either Scheme I or Scheme II (see above). The lattice summation was done by choosing a reference molecule and calculating pairwise interactions with increasingly distant neighbors until the magnitude of the interactionbetween a pair of moleculeswas less than 0.02 kcal mol-‘. The kinetic energyof the lattice vibrationsELAT was calculatedfrom experimental and/or calculated lattice frequencies. The kinetic energy of optical modes was calculated according to the Einstein app~ximation [41]. The kinetic energy of acoustical modes was calculated according to the Debye approximation [41]. The lowest optical frequency was taken as the maximum frequencyvMAXin the Debye formulation. For a crystal whose geometryis known at a given temperaturefrom crystallographic data, determination of E PACKand EMT allows the calculation of A.&un from eqn. ( 10). The calculatedvalue of AHsunmay be compared with the experimentalenthalpyof sublimation.By usingeitherSchemeI or Scheme II in the dispersion energy contribution to E PACK,it is possible to determine which scheme is preferable (see below). Ilet&
of the electrostatic and dispersion seems
One molecule (5-HIND, 5-HT or 6-HIND) was kept fixed while the scanning molecule (indole) was oriented with its molecularplane parallel to that of the fixed molecule at an interplanardistance of 6.238 bohr. The indole portion of the fmed molecule was taken to be the xy plane with the origin at atom Nl of the indole ring. The y axis was directed along the Nl-C2 bond. The coordinate system is depicted in Fig. 1. The scanning moleculewas translated
q -7.0 0 -5.0 A -3.0
7
A I
I
-10.0
-5.0
5.0
1
I
I
0.0
10.0
15.0
X (bohr) a
s 0
ui 0
E
d
0
e >
0
Y 0
s
A
0 .
0
? I
-10.0
-5.0
I
0.0
I
I
5.0
10.0
1
15.0
X (bohr)
Fig. 1. Total interaction energy ICecaNbetween a fixed molecule (shaded) and a scanning molecule is calculated as the sum of electrostatic and dispersion energies, and is represented as isoenergy contours (kcal mol-I). The scanning molecule is in a parallel plane 6.238 bohr from the fixed molecule. Sketches of the scanning molecule are shown at positions corresponding to local energy minima. A. 5-HIND (fixed) and indole. B. 6-HIND (fixed) and indole.
along a square grid with a step size equal to 0.5 bohr. At each point, the indole was rotated counterclockwise in increments of 10” about an axis perpendicular to the molecular plane through the midpoint of the C&C9 bond (the position where the pyrrole and benzene rings are fused). At each point, values of Eas, EDIsp, the lowest interaction energy and the corresponding orientation angle were saved. Once the lowest energy had been determined for each point on the
329
grid,positions of local energyminima were found by comparison of the energy at a givenpoint with the energyat neighboringpoints. The interactionenergy for the entire scan was representedby means of isoenergycontour maps. The molecules5-HIND, 6-HIND and indole each have symmetryC,. Thus, scans of indole with 5-HIND or 6-HIND can have two distinct relativeorientations. In the “P” scan, the initial o~en~tion of the scanning indole superimposes it with the indole portion of either B-HIND or 6-HIND so that the angle between the respective C&C9 bonds is zero (scan angle, 0” ). In this orientation,the calculateddirection of the dipole vector of the scanningindole is 195’ with respectto the x axis. In the “M” scan, the initialindole orientation (scan angle, 0” ) was generatedby reflecting the starting orientation for the “P” scan through the yz plane which passes through the Nl-C2 bond. As a resultof this reflection,the correspondingdirection of the indole dipole is 345’ with respect to the x axis (see Fig. 1) . Since the side-chain of 5-HT is out of plane, the molecularsymmetryis C,. Thus, there are four possible scans of indole relativeto 5-HT. To avoid close contact between atoms of the two molecules,the only scans actuallyrun were the “P” and “M” scans with the indole on the side of the plane distal to the side-chain. RESULTS AND DISCUSSION
Comparison between calculated and e~~ri~ntal
AHsUB
The comparison between calculated and experimentalenthalpies of sublimation for benzene and naphthalenecrystalsis shown in Table 2. For benzene, the value of AHsus has been measuredat 273 K [ 421, but high resolutionneutron diffraction data [ 241, used to calculateAHsUB,have been measuredat 138 K and 218 K. Thus, calculated values of AHsoa at the last two temperatures were linearly extrapolated to 273 K so that comparison could be made with experiment. A similar extrapolative method was employed by Banerjee and Salem [43]. In the calculation of EMT (see the Meth~ section), fr~encies of lattice vibrationsfor benzene at 138 K [ 42 ] and naphth~ene at 298 K 144] wereused, Since experimentallattice frequencieswere not known for benzene at 218 K, the value of EUT was estimated from a linear regressionof ELATversus temperature.For several molecular crystals, previously reported lattice frequencies were used to calculate values of ELAT.The crystals used, along with the correspondingtemperatures,were benzene at 138 K [ 421, naphthaleneat 298 K [44 1, cu-p-C,H&l, at 300 K [41], /3-p-CGH4Clz at 318 K [41], pyraxine at 180 K and 300 K [42], a-s-triazine at 300 K [42], j&&iaxine at 150 K [42] and s-tetraxineat 253 K [ 421. The crystalslisted are composed of neutralnon-
polar aromatic’molecules of similar size. The equation for the linear regression of ELAT versus temperature (TEMP ) is E LAT=co +c,(TEMP)
(12)
A fit of the nine data points to this equation yields the coefficients c,=O.154 kcal mol-’ and c,=O.O115 kcal mol-’ K-l. The points are closely correlated (r= 0.999; P< 0.~1) - From this regression, the predicted value of EMT for a crystal at 218 K is 2.7 kcal molW1. As expected for neutral non-polar molecules, the magnitudes of EsCF(xl ) for benzene and naphthalene shown in Table 2 are small in comparison with the corresponding magnitudes of EnIsp (xl). Thus, EPAcx depends primarily on dispersion. Neutron diffraction structures for benzene [ 241 at 138 K and 218 K and X-ray diffraction structures for naphthalene [ 251 at room temperature (taken to be 298 K) were used in the calculation of dispersion energies. The TABLE 2
Comparison between calculated and experimental values of enthalpy of sublimation for benzene and napbthalene’ Molecule
138 1.1 1.7 0.4
T WI 4RT=EGAsSPV E b EZ(xl)C
218 1.7 2.7 -0.4
273
296 2.4 3.6 1.1
Schemed
EnIsP(xW
E PACKf W”B (talc 1s A&&XptP
I
II
I
II
- 16.4 - 16.0 15.4
- 13.4 - 13.0 12.4
- 14.9 - 15.3 14.3
-12.2 - 12.6 11.6
II
I -
-
13.5 10.7
11.1
I
II
-23.5 -22.4 21.2
- 19.0 -17.9 16.7 17.3
*Energies in kcal mol-‘. bKinetic energy of lattice cakulated from frequencies of lattice vibrations as described in text. Frequencies for benzene at 138 K from ref. 42. Erequencies for naphthalene from ref. 44. The vahre of .8&T for benzene at 218 K wss estimated from a linear regression (eqn. (12)). “SCF contribution to lattice potential energy corrected for BSSE (ref. 33). dScheme I: cyTfor C-CAROM= 0.59 A3. Scheme II: aT for C-Cmo,=0.40 A3 (see text). “Dispersion contribution to lattice potential energy calculated using eqn. (5). Total lattice potential energy calculated using eqn. (11). pEnthalpy of sublimation calculated using eqn. (10). Values for benzene at 273 K were obtained by liuear extrapolation from corresponding values at 138 K and 218 K. hExperimental enthalpy of sublimation. Value for benzene from ref. 42; value for naphthalene from ref. 45.
331
bond polarizabilitiesfor C-H bonds were taken as cyL=arT=0.64 A”. For aromatic C-C bonds, (Yewas taken as 2.21 A3and aT was taken as either 0.59 A” (Scheme I) or 0.40 A3(Scheme II). The mean excitation energy U for each moleculewas set equal to 1.8 times the experimentalverticalionization potential (see Table 1) . Thus, UBmz= 16.63 eV and UN-n = 14.67 eV. In each of the cases treated,Emsp(xl) makes the predominantcontributionto EPAcx.In fact, B&-&xl) could have been omitted from the collations ~thout introducing significant errors into the comparison between the calculated and experimentalvaluesof AI&n. It maybe seenin Table 2 that valuesof Msua calculatedaccordingto Scheme II ( aT for C-CAROM=0.40 A3)arein better agreementwith experiment for both benzene [ 421 and naphthalene [ 451. Consequently,Scheme II was used in all of the dispersion energycalculationsreportedbelow. ~~c~ost~t~ and dispersion stuns
Isoenergycontour maps of EscANfor the interactionof indole with 5-HIND (“P” scan), and 6-HIND (“P” scan) are presentedin Fig. 1. Parametersused TABN
3
Parameters used in the calculation of the dispersion energy Molecule
Mean excitation energy“ (kcal mol-‘)
Benzene Napht~ene Indole B-HIND 6-HIND 5-HT
383.64b 3383Ob 327.92” 326.2Sd 317.13d 321.28d
Bond
Bond polarixabilit~ (A3) Longitudii
C-H c-c C-GEi,,’
C-N c-o N-H O-H
0.64 0.99 2.24 0.57 0.89 0.50 0.58
‘1.8 times vertical ionization potential. bVertical ionization potential from ref. 38. cVertical ionization potential from ref. 36. dvertical ionization potential from linear regression (see Table 1) . “VaIues for O-H bond from ref. 40. Other values from ref. 39, ‘All bonds between heavy atoms in indole ring taken as C-C (arom). @I’rausverse~~~b~ity taken as mean of in-plene and out-of-phme vahres.
Transverse 0.64 0.27 0.408 0.69 0.46 0.83 0.79
332 TABLE 4 Positions, energies and orientations of local energy minima derived from electrostatic and dispersion scans with indole” “Fixed” molecule Scan type Position of minimumb Eae 5-HIND
“P”
“M”
B-HIND
“P”
5-HT
=P*,
( - 1.5, -3.5) ( 6.5, 6.5) ( 7.0, -7.0) ( -3.5, 0.0) (-1.5, -4.0) ( 5.5, 6.5) ( 6.5, -7.0) ( -2.0, -3.0) ( 11.0, -0.5) ( - 1.5, -3.5) ( 7.0, -7.0) ( 7.0, 6.5)
-3.56 -2.20 -3.72 -3.74 -3.59 -2.69 -3.47 -3.94 -3.69 -3.72 -3.69 -2.87
EDrsp EsCAN Dipole angle” (deg) -4.43 -3.62 -3.67 -3.79 -4.07 -3.70 -3.81 -4.23 -3.79 -4.48 -3.67 -4.72
-7.99 -5.82 -7.39 -7.53 -7.66 -6.39 -7.28 -8.17 -7.48 -8.20 -7.36 -7.59
25 265 95 25 55 265 115 15 175 25 95 255
“Positions in bohr, energies in kcal mol-‘, orientations in degrees. bx-y coordinates of midpoint of C&C9 bond in scanning indole. ““P”scan: (dipole angle) = 195” + (scan angle); “M”ecan: (dipoleangle) = 345” + (scan angle).
to calculateEDIsPfor these and other scans are given in Table 3. A sketch of the fixed molecule is shown in each map along with a sketch showingthe position and’orientation of the indole at each local energy minimum. Table 4 gives the position and orientation of each local energy minimum along with the values of EscAN,Eoo and EnIsp.In addition to data for the scans shown in Fig. 1, Table 4 also contains data for two other scans (5 HIND “M” and 5-HT “P”). In comparing the two scans involving &HIND, one observes that for each local rni~~urn in the “P” scan there is a correspondingminims in the “M” scan with a similarposition and energy.In addition,the dipoles of a pair of correspondingminima (one “P”, one “M”) have similarorientations.Similar trends are observed when comparingthe “P” and “M” scans for 5HT or 6-HIND (data not shown). The position, orientationand energyof each local minimumin the 5-HIND scan closely correspond to a minimum in the 5-HT scan. This finding thus supportsthe use of 5-HIND as a model for 5HT, especiallysince the scan of 5-HT with indole was done on the side of the XYplane away from the 5-HT side-chain. The “P” scan with 5-HIND is quite different from that with 6-HIND (see Fig. 1) . Only the minimanear the pyrroleportion of the furedmoleculesare in close agreement.Each map is characterizedby a region of weak interaction which spans the indole ring and the hydroxyl group. The difference in the positions of the O-H group leads to a large difference in the characterof the contour maps. When the scanning molecule is close to the oxygen atom, the
333
magnitude of EscAN is small. As a result of the pronounced effect of the oxygen atom on the shape of the contour map, the position of the local minimum at (7.0, - 7.0) in the &HIND scan corresponds to a position of very weak interaction in 6-HIND. For this minimum in the &HIND scan, the value of .&CAN is - 7.39 keal mol- l at an orientation angle of 260’. For the same position in the 6-HIND scan, the value of EscAN is -1.61 kcal mol-’ at a similar orientation angle of 290”. This difference of 5.78 kcal mol-” in the interaction energy comes p~m~ily from the electrostatic infraction. For 5-HIND, EQa is -3.72 kcal mol-’ and Emsp is -3.67 kcal mol-l. The corresponding values for the 6-HIND are + 2.45 kcal mol-* and -4.06 kcal mol-I, respectively. Thus, the difference in EQa is responsible for the selectivity demonstrated by the model of a neutral recognition site in the receptor. If one assumes that the geometry of B-HIND at the local minimum identified by the scan corresponds to a high-affmity binding configuration of 5HT at a 5-HT receptor, then 6HT would be expected to have a much lower affinity for that receptor because it will have an ~favor~le electros~tic interaction with the receptor site. In fact, experimental measurements show a large difference in affinities between 5-HT and 6-HT at both 5-HTlA and 5-HT2 receptors. Binding studies at 5 HTXAreceptors [46] give dissociation constants of 3.1 nM for 5HT and 1600 nM for 6-HT. According to measurements of 5HTz receptors in the isolated rabbit aorta [47], the dissociation constant for 5HT is 250 nM, whereas the dissociation constant for 6-NT is greater than 10 000 nM,
Since the scanning position at (7.0, - 7.0 ) clearly distinguishes between 5-HIND and 6-HIND, the configurat,ion from the “P” scan of 5HIND with indole at that position was selected for a de~led investigation of the variation in scan energies with the orientation angle. The results are shown in Fig. 2. The magnitude of E nIsP is generally greater than that of I&o, and the variations in Eoo and Enrsp are almost exactly out of phase. The most favorable total infraction energy occurs at an orien~tion angle of 260” between the C& C9 bonds of the two molecules. In this orientation, both the electrostatic and dispersion terms are attractive and contribute equally to the stabilization of the complex. Since the two molecules do not overlap appr~iably (see Fig. 1A ) , the electrostatic interaction is at a minimum and the dispersion interaction is at a maximum. On the other hand, maximum overlap of the two rings (which occurs at an orientation angle of 150” between the (X-C9 bonds) gives rise to a repulsive electrostatic energy and an attra~ive dispersion energy (see Fig, 2 ). The angular variation in the total interaction energy, EscA, follows that of IZoo because Eee varies over a range of approximately 4.8 kcal mol-‘, while EnIsp varies only over a range of approxima~ly 1.5kcal mol-“. Consequently,
334
0
ELEC
A DISP
404 0
I
I
I
I
I
60
120
180
240
300
ORIENTATION
ANGLE
I 360
(degrees)
Fig. 2. Dependence of interaction energies between &HIND and indole on the orientation angle between the C&C9 bonds of the two molecules. The relative position of the two molecules is at local minimum (7.0, -7.0) as shown in Fig. IA. ELEC, electrostatic energy; DISP, dispersion energy; TOTAL, total interaction energy.
the electrostaticinteractionis the importantproperty in determiningthe preferred orientation and in establishing a molecular correlate of selectivity, whereasthe dispersion energyprovides the major portion of the stabilization of the complex. Hunter and Sanders [48] reached a similar conclusion in a recent analysisof 7c-xinteractionsin porphyrins. Similar orientations properties were found in scans in which i~~olium was used as a model of a receptor that contains a chargedresiduein it [f&10]. However, the mechanismsfor selectivity and affinity seem to be different in the two models. In the chargedmodel both the interactionenergy,a correlate of affinity, and the orientation of the interactingmolecules, a basis for selectivity, were determinedby electrostatics. In the neutral model, which is explored in this work, the electrostaticinteraction governsthe orientationalselectivitywhereasthe dispersioninteractioncontributesmainly to the affinity. Furthermore,in the charged model the orientational selectivity was determined by the favorable interaction between the ~stribution of the positive charge in irni~o~~ and the electros~tic potential of 5HT, whereas the interactionbetween the neutralindole and 5-HT may be better characterized by the interactionbetween the indole dipole and the electric field of 5-HT. The interactionenergyEWbetweenthe indole dipole p and the 5-HT electric field Eis givenby Ep=
-~-ST
(13)
Because the electric field Eis the negative gradient of the molecularelectrostatic potential of 5-HT, Fig. 3 showsthe electrostaticpotential of 5-HT in the
335
IJ -16.0 O -12.0 A -8.00 + -4.00
0
+ 0
:
s 0 -10.0
-5.0
0.0
5.0
10.0
15.0
f;; *
20.0
X (bohr)
Fig. 3. Dipole vectors of indole at local minima obtained from a scan of 5-HT and indole are superimposed on the molecular electrostatic potential map of 5-HT calculated in a plane 6.238 bohr from the plane of the indole portion (see the Methods section). The direction of each dipole vector is approximately aligned with the electric field generated by 5-HT, which is perpendicular to the equipotential contours (kcal mol-’ ) .
plane (z= -6.238 bohr ) in which the electrostaticand dispersion scans were carried out. Sketches of the local minima for indole obtained from the 5HT “P” scan are superimposedon the electrostaticpotential map. For each indole, the dipole vector is shown at the midpoint of the bond between C8 and C9. It may be seen that the direction of each dipole vector is approximatelythe same as the direction of the electric field of 5-HT, which is perpendicularto the contours. At each local minimum,the indole molecule is located in a region in which the electric field is relativelystrong and uniform, i.e. a region where the equipotential contours are closely spaced and approximatelystraight. Thus, the dipole-electric field interaction model may be used to interpret the electrostatic and dispersionscans in Fig. 1. Stronginteractionsoccur in regionswhere 5-HIND or 6-HIND has a strongelectricfield. At (7.0, - 7.0), a position where the interaction energywith indole clearly distinguishesbetween &HIND and 6-HIND, the electricfield of 5-HIND is strongbut the field of 6-HIND is weak. Interaction energies of 5-HIND complexes
The calculated interaction energiesfor the complex of B-HIND and indole at the (7.0, -7.0) position from the “P? scan are presented in Table 5. The interplanardistance is 6.238 bohr.
336 TABLE 5 Camparison of interaction energies’ in &HIND complexesb B-HIND and indole “P” (7.0, -7.0)
&HIND and imidazolium (7.5,3.0)
-3.72 -3.67
-11.97
- 7.39
&C.&N=&QfhSF’
SCE Morokuma decomposition -11.86 1.62 -0.94
ES EX
- 3.90 1.22
PL CT MIX
-0.35 -0.23 0.02
=%m
- 3.24
- 11.61
&xxfco~)
-2.70
-11.11
&wr~t”&w
(corr) +EDISP
-0.46 0.03
-6.31
*Energies in kcal mol-‘. ‘~~~~ distance is 6.238 hohr, cEm~ could not be c&&ted for ~i~~li~ ities af charged molec&~ are not available.
complex because parameters for bond polarizabil-
accordingto the scheme The SCF ~bi~tion energy,EscFwas diurnal of Morokuma [30,31] . The value of E s& was corrected for basis set superposition error [ 331 to givea quantitydesignatedas EeCF(corr). Interestingly,the valueof je,, is in close agreementwiththe valueof the electrostaticterm (ES) a Thus, at this interplanardistance, the monopole-monopole approximationof the electrostaticinteraction gives a reasonable representationof the electrostatic energy. The EB1sPaccounts for 58% of EmYAL,the total s~ili~ation energy.Thus, tbe s~b~ity of the complex between &HIND and indole has a ~onsiderahle~ontributioufrom the dispersion in~~~~ion. It is of interest to compare the Moro~ma decomposition of the energy of i~~ra~tiou in the complex of G-HINDand indole with that in the complex of &HIND and imidazolium cation (see Table 5). Indole represents a neutral model for a 5-HT receptor,whereasimidazoliumrepresentsa chargedmodel. The local minimumconfigurationsof the 5-HIND-imidazolium complex were determinedfrom a mo~opole-mo~opole scan, using LP-3G charges,at an interplanardistance of 6.236 bohr. The local minimumat position (?.S, 3.0) in the coordinate system used for the ~-HIP-~dole scans correspmds to that
337
obtained previously in similar scans of B-HT with imidazolium [ 101. Similar to the alignment of the dipole vector of the scanning indole with the electric field of 5-HT (see Fig. 3), we find that the direction of the dipole vector of imidazolium also coincides with the direction of the electric field of &HIND. The most important difference between the two complexes is in the SCF interaction energies. In the complex with imidazolium, EsCF is - 11.61 kcal mol-’ whereas in the complex with indole it is only -3.24 kcal mol-‘. The decomposition of the interaction energy in the 6-HIND-imidazolium complex provides an explanation for this large difference. The magnitude of ES in the 5HIND-imi~zolium complex ( - 11.86 kcal mol-‘1 is much larger than the correspon~ng value in the 5-HID-indole complex ( - 3.90 kcal mol-l ). The ma~i~des of EX (1.62 kcal mol-” 1, PL ( - 0.94 kcal mol-l ) and CT ( -0.46 kcal mol-’ ) are similar to the correspondingvalues for the indole complex (see Table 5). We also note that although the decomposition of EsCF for the two complexes is quite different, the BSSE is approximately 0.5 kcal mol- ’ for each complex. Thus, it appears that the BSSE reflects the size of the molecular basis set rather than the nature of the infraction between &HIND and the respective receptor models. In the imidazolium complex, the contribution of the electrostatic term to the SCF energy is the main stabilizing factor, whereas in the indole complex dispersion plays an important role, contributing over 50% to the interaction energy. The methods used here to calculate dispersion energies could not be applied to the imidazolium complex because parameters for bond polarizabilities of charged molecules are not available. However, the polar~bi~ty of the imidazolium cation is expected to be small, and ~onse~ently the dispersion interaction energy of the imidazolium complex is expected to be less than that of the indole complex.
The changes in electron density upon formation of the complex between 5HIND and indole at the (7.0, - 7.0) position from the “P” scan were studied in two related ways. Values of Ap for the complex with Az equal to 6.238 bohr, corrected for BSSE and calculated by integration in planes parallel to the molecule planes (see the Methods section), are plotted in Fig. 4. The figure shows a depletion of electron density midway between the indole planes. This probably arises from repulsion between the electron clouds of S-HIND and indole. Simul~eously, the electrons in each molecule are polarized in a direction perpendicular to the indole plane, so that the negative end of each induced dipole points toward the region between the planes. In this manner, the electron charge of each molecule of the complex is polarized towards the other molecule. It may be seen in Fig. 4 that the polarization of 5-HIND is slightly greater than that of indole. The
INDOLE
4.23*
DISTANCE
S-HIND
0.000
(bohr)
Fig. 4. Electron density (e/bohr X 103) redistribution in the (B-HIND + indole) complex at local minimum (7.0, - 7.0) (see Fig. 1A) calculated from molecular wavefunctions obtained with the LP-3G basis set. 4p is calculated by integration in planes parallel to the molecules in the complex and~~~ed~~(~mplex)-~(5-H~+gh~t ~dole)-~(indole+gh~t 5-HIND). See the Methods section for details.
overall pattern of the charge redistributionexplains why dispersion,which is a correlated interaction between induced dipoles, makes a large contribution to the total stabilizationenergyof the complex (see Table 5). The electron density redistributionin the complex of S-HIND and indole contrasts sharply with that in the complex of 5-HT and imidazoliumcation which was reported previously [ 81. In the latter case, 5-HT was more extensively polarized (toward the imi~oli~) and the imi~oli~ was slightly polarized away from the 5-HT. In the ~i~zoli~ complex, 5HT was polarized nearly eight times more than 5-HIND in the indole complex. The depletion of electron density midway between the molecule planes was relatively small.The polarizationpattern in the 5-HT-imidazolium complex was shown to be characteristic of a strong electrostatic interaction. The difference between the polarization in the imidazolium and indole complexes is also reflected in the difference between the PL terms in the corresponding energy decompositions (see Table 5). A more detailedanalysisof the electron density redistributionwas obtained by computingdifference maps in planes parallelto the indole planes. Figure5 shows maps for the planes at a distance of 1.9 bohr on either side of each molecule. Thus, Figs. 5A and 5D represent charge redistributionsin planes outside the molecularcomplex and Figs. 5B and 5C) in planes inside the molecular complex. Each of the selected planes corresponds approximatelyto a a-electron region in one of the molecules.In all four planes, there is a distinct polarization of electron density away from the region in which the molecules overlap.This polarizationis moreextensivein the spacebetweenthe molecules as can be seen from a comparison of Figs. 5B and 5C with Figs. 5A and 5D.
339
0
0
vi-
vi
0
0
c d
d
‘>
9
T
0
80
*
?_
Lp
‘f 0
s-
q
0
-0.30
_
0
-0.30
s D s
+ -10.0
-5.0
0
A
0.30 I
I
0.0
I
5.0
I
10.0
I
?
15.0
X (bohr)
B -10.0
I
-5.0
I
0.0
I
5.0
I
10.0
/
15.0
X (bohr) d
X (bohr)
X (bohr)
Fig. 5. Electron density (e/bow X 104) redistribution in the (5-HIND + indole) complex at local minimum (7.0, - 7.0) (see Fig. 1A) calculated from molecular wavefunctions obtained with the LP-3G basis set in planes parallel to the indole portions of the molecules. (B-HIND in plane z = 0; indole in plane z = - 6.238 bohr ). Shaded regions correspond to increases, and unshaded regions to decreases in electron density upon complex formation. Each selected plane corresponds approximately to a x-electron region in one of the molecules. A. z = + 1.900 bohr. B. z = - 1.900 bohr. C. z= -4.338 bohr. D. z= -8.138 bohr.
This charge redistribution reduces the repulsion between the electron clouds of the two molecules, and thus stabilizes the complex by inducing oppositely directed dipoles parallel to the planes of the two molecules (compare Fig. 5A with Fig. 5D and Fig. 5B with Fig. 5C). The formation of these induced dipoles gives additional support for the importance of polarizability in the interaction complex between 5-HIND and indole.
CONCLUSIONS
We haveexplored the possibility that the recognitionof 5-HT by two different receptors is governed by different physical interactions. It was demonstratedthat 58% of the interactionenergyof 5-HT with a neutralligand,modeled by the indole portion of tryptophan, was due to dispersion;the rest was mostly due to elect~static infractions. This is in clear contrast to the previously studiedinteraction of 5-HT with imidazoliumcation [ 8-10 1, where the’ interaction was found to be primarily electrostatic. In spite of the different interactionsthat contributeto the stabilityof the complexes,the most important element in the recognition of 5-HT by both of these receptor models is electrostatic.With indole servingas the model, the recognition is based on its polarity, and is expressedin the interactionof its dipole with the electric field of &NT. With imidazolium,on the other hand, the recognitiondepends upon the interactionbetween the distributionof its positive chargeand the electrostatic potential of 5-HT. Assessmentof the roles that these models (indole and imidazo~um) play in 5-HT receptorsrequiresadditionalexperimentalinformationabout the structures of the receptorproteins. In the meantime,we can determinewhetherthe demonstratedselectivityof differentdrugstowardsthe known 5-HT receptors can be correlatedwith the distinguishingproperties of the models. The different mechanismsthat are responsible for affinity-dispersion in the neutral model and electrostatic in the charged model-illustrate a possible basis for the specificityof ligandstowards the various 5-HT receptors. ACKNOWLEDGMENT
We thank Dr. Hare1Weinsteinfor his helpful comments on the manuscript. Computations were performed on the supercomputersystem at the Cornell National SupercomputerFacilitywhich is sponsored by the National Science Foundation.The authorsgratefullyacknowledgethis resourceand the generous allocationsof computertime from the UniversityComputingCenterof the City Universityof New York. This work was supported by the National Institute on Drug Abuse under grant DA-01875 and by the National Institute of General Medical Sciences under grant GM-34852. REFERENCES 1 2 3
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