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PMR conformational studies of Pd(II) complexes with Ala-Tyr and d -Leu-Tyr depeptides
Journal of Molecular Structure, 50 (1978)
73-80 o Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands
PMR CONFORMATIONAL STUDIES OF Pd(I1) COMPLEXES WITH ALATYR AND D-LEU-TYR DIPEPTJDES
HENRYK
KOZ%OWSKI,
Institute of Chemistry, (Received
MALGORZATA
JEZOWSKA and HELENA SZYSZUK
University of Wrocibw, Joliot-Curie 14, 50-383,
Wroctaw (Poland)
13 March 1978)
ABSTRACT
PMR studies of Pd(II) complexes with Ala-Tyr and D-Lcu-Tyr role of metal ion in inducing tridentate
coordination
the dipeptide
of dipeptide
conformation
(pH 3-10)
revealed the essential in solution. In a complex with
the most stable conformer
residue is that with the aromatic ring in gauche position
to carbgxyl
of a
tyrosine
and amide groups. At
is a bidentate ligand (NH,, N-), the tyrosine residue changes its conformation drastically, and the most stable conformer is that with the aromatic ring in trans position to the carboxyl group. It was found that the C-terminal amino acid does not have
high pH a dipeptide
any considerable
influence
on the conformation
of an N-terminal
amino acid in both kinds
complexes with palladium(D). Increasing the temperature up to 370 K has only a minor effect on the conformer population in solution. of
INTRODUCTION Our recent studies on the Pd(II) complexes with dipeptides containing the aromatic ring in an amino-acid side chain [ 1,2] revealed that the metal ion induced some favourable conformation of such a side chain which varied considerably compared to that found in a metal-free ligand. For the studied complexes, i.e. Pd( II) with glycyl-L-tyrosine (Gly-Tyr), glycyl-L-phenylalanine
(Gly-Phe) and L-phenylalanyl-glycine (Phe-Gly), in the most stable conformer the aromatic ring was above the square planar complex plane. To explain the stabilization of such a rotational isomer (gauche) an interaction between aromatic ring and metal ion has been proposed [ 1,2] . X-ray studies of some Cu( II) complexes with tyrosine [ 3-63 and tryptophane [ 71 containing ligands have shown that the interaction between the metal ion and side-chain group was possible but it was not the rule [5] . The molecules which have the interaction between aromatic ring and copper (II) ion are in the same approximate conformation as that in the Cu(I1) complex with L-leucyl-L-tyrosine (no interaction) [5] . Thus, the conformation of the amino-acid side chain would not be a good criterion for determining whether an interaction will occur. In Pd(I1) complexes, however, such interaction happens more easily because there is no apical coordination to the metal ion, which is the case in cupric complexes [3-G].
74 EXPERIMENTAL Glycyl-L-tyrosine ( Gly-Tyr), L-alanyl-L-tyrosine and D-leucyl-L-tyrosine (D-Leu-Tyr) dipeptides were used as obtained from Fluka AG. K2PdC!14was obtained by crystallization of KC1 and PdClz solutions containing HCl. Analysis showed a purity better than 99%. PMR spectra were recorded on a JEOL 100 MHz JNM PS-100 spectrometer using tert-butyl alcohol as an internal standard. All spectra were measured at 25 * 2”C, and those for Pd(II)Gly-Tyr solutions were measured up to 98°C. The pH was adjusted on a MeraElmat N-5122 pH-meter. PMR spectra of tyrosine oCH$CH2 protons have been treated as of ABX type and they were analyzed on a JEC-6 computer. The concentration of dipeptides was 0.15 mole 1-l in all samples. DzO (99.8%) was used as a solvent. RESULTS The PMR parameters for metal-free Ala-Tyr and D-LeU-TYr at chosen pH are listed in Tables 1 and 2. The rotational isomer notation for tyrosine residue is given in Fig. 1. The fractional population of each isomer can be estimated by the Feeney [8] or Pachler [9] approximation (Tables 1 and 2). The data TABLE 1 ‘H NMR parameters and rotamer populations for Ala-Tyr dipeptide and its complexes with Pd(II) ion (the populations obtained by Feeney’s equations are given in parentheses) PH
VA
vx
iJAB
JAX
JBX
WH@
VCH
0.53 175.2
191.3
336.4
13.9
9.0 5.7
24.8
276.8
2.69 173.2
189.2
331.2
14.0
8.5 5.4
24.6
273.8
5.64
164.8
184.8
311.9
14.0
9.0 4.9
24.5
271.3
8.47
162.8
185.8
312.7
14.0
8.9 4.8
3.9
236.8
10.32 158.7
182.0
311.2
14.0
8.9 4.8
-8.9
213.8
13.40 156.9
177.4
309.4
14.0
8.8 4.6
-7.8
214.6
Pd(il)-Ala-TyrI:1 8.60 167.9 197.9
300.2
13.7
2.9
5.7
4.0
230.0
13.80 161.0
166.5
297.3
13.8
11.8
3.9
-22.3
Pd(II)-Ala-TyrI:2 13.59 133.6 177.7
315.6
13.9
11.5
2.9
-16.1
Pl
Pa
0.58 0.28 (0.64) (0.36) 0.54 0.25 (0.59) (0.40) 0.58 0.21 (0.65) (0.27) 0.57 0.20 (0.64) (0.25) 0.57 0.20 (0.64) (0.25) 0.56 0.18 (0.64) (0.22)
P3 0.14 (0.00) 0.21 (0.01) 0.21 (0.08) 0.23 (0.11) 0.23 (0.11) 0.26 (0.14)
0.03 0.28 0.69 (0.00) (0.12) (0.88) 0.04 218.1 0.84 0.12 (0.97) (0.03) (0.00) 228.2
0.81 (0.96)
0.03 (0.04)
0.16 (0.00)
75 TABLE
2
‘H NMR parameters and rotamer populations for D-Leu-Y&r dipeptide and its complexes with Pd(II) ion
PH
“B
“X
IJAB
JAX
4.04 149.5
205.8
332.0
14.3
12.0 4.0 -51.7
254.7
6.08
148.4
204.4
326.4
14.1
12.0 3.8 -52.9
255.4
9.78
149.1
195.3
324.2
14.2
10.9
4.2
-53.3
200.5
11.62
148.3
183.2
315.4
14.1
9.6
4.2
-48.4
203.5
13.50
148.1
181.2
314.0
14.1
9.8
4.2
-47.4
203.9
Pd(Il)--o-Leu-Tyr 1:1 8.20 164.0 208.6
294.9
13.5
2.8
5.4
-30.4
225.5
13.90
280.0
14.4
-33.4
184.8
11.3 3.3
-28.5
200.1
“A
169.4
Pd(ZI)-D-Leu-Tyr
13.76 133.2
JBX
UCH,h”
“CH
Pl
P2
PO
0.02 (0.00) 0.04 (0.00) 0.14 (0.13) 0.14 (0.19) 0.14 (0.18)
0.11 (0.00) 0.22 (0.08) 0.21 (0.07)
0.02 0.25 (0.00) (0.07) 0.84 (0.92)
0.73 (0.93) 0.16 (0.08)
0.79 (0.93)
0.15 (0.00)
0.75 (0.87) 0.64 (0.73) 0.65 (0.75)
1-2
170.3
323.1
13.7
0.06 (0.07)
obtained are in good agreement with those given by Dale and Jones [lo] for both peptides. ln tyrosine residue, over the whole pH range, the most stable rotamer is that with the carboxyl group and aromatic ring truns to each other (1). The gauche isomer (3) is rather unstable in metal-free peptide, though its population in basic solution is quite considerable at least for the Pachler approximation (Tables 1 and 2). Pd(ll)
: dipeptide
1: 1 molar solutions
Addition of Pd(I1) ions to a dipeptide containing solution changes the PMR spectra of the latter dramatically. For 1: 1 solutions in pH range 3-10, the PMR spectra have the parameters given at pH 8.6 for Pd(II)-Ala-Tyr solutions (Table 1) and at pH 8.20 for Pd(II)-&Leu-Tyr ones (Table 2). The chemical shifts as well as the coupling constant variations in comparison to the metalfree ligand are caused by the metal ion coordination to the dipeptide molecule. The possible coordination sites of Pd(II) ion in both peptides are the amine group, the deprotonated nitrogen of a peptide linkage and the carboxyl group. At any pH value between 3 and 10 all these three donor groups are coordinated to the metal ion as has already been found for similar systems [l, 2,11,12, 13) . Also d-d transition energies found for both kinds of solutions, i.e. 332 and 333 nm for Pd(II)--Ala-Tyr and Pd(If)-D-Leu-Tyr, respectively, may suggest the trident&e coordination by NHP, N- and COO- donors [2,11,13].
76
The conformation of a tyrosine residue of those tridentately coordinated dipeptides is quite different to that found in metal-free ligands (Tables 1 and 2). There is a considerable increase of the rotamer 3 (gauche) population, and the rotamer 1, most stable in the metal-free ligands, is almost absent in the complex solution (p I <0.05). An increase of pH above 13 in 1:l Pd( II)-Ala-Tyr solutions leads to the hydrolysis of metal-car-boxy1 bonds [ 1, 2, 11, 12,133 , which results in quite drastic changes in the PMR spectrum (Table 2). There is an upfield chemical shift of all protons of dipeptide molecules except aCH-Tyr (Table 1). There is also a large increase in JAx value (2.9 to 11.8) and, as a consequence, a large increase of rotamer 1 population (-O-t- 0.9). The rotamer 2, which was quite appreciably populated for tridentately coordinated and metal-free Ala-Tyr, vanished almost completely (pl = 0.02; Table 1). Similar variations in the PMR spectrum were found for the 1 :l Pd( 11)-DLeu-Tyr solution above pH 13. The spectrum of the tyrosine residue, however, is of A2X type (or rather deceptively simple ABX) and it is not possible to distinguish the contributions of rotamers 1 and 2. In both cases, the decrease in gauche rotamer population was either to the same or to an even lower value than that was found for the metal-free ligand in the same pH region (Tables 1 and 2). Pd(II)
: dipeptide
1:2 molar solutions
The PMR spectra of 1:2 metal to peptide molar solutions up to pH 12 consist of two sets of lines, one for the 1:l complex described above and the other for metal-free ligand. Pd(II)-peptide systems are quite inert on an NMR time-scale and we usually observe a separate PMR spectrum for each chemical species in solution. Above pH I.3 only one single spectrum is observed, suggesting the coordination of two dipeptide molecules to a metal ion. A break of the metalcarboxyl bond makes the formation of two chelates with two ligand molecules coordinated by NH2 and N- possible [l, 2,121. Coordination of four nitrogen donors to Pd(I1) ion is also supported by the absorption spectra. The d-d transition energies (h <300 nm) found for 1:2 Pd(II):Ala-Tyr and Pd(II):D-Leu-Tyr are close to the values found for Pd(II) ion with four coordinated nitrogens [Z, 13,14,15]. An overlapping of the d-d transition band with that for WV* in the aromatic ring (Tyr) does not permit the precise calculation of the former transition energy. DISCUSSfON X-ray studies have shown that the metal ions coordinated to a peptide molecule do not affect considerably the dihedral angles in tyrosine residue [3-6,16] , which were assumed to be 60”, 180” and 300” (Fig. l), and that in coordinated or metal-free ligands these angles could deviate by about 3-8”
77
CO’JH I
W-kOH COOH &HL OH
H0 d H;N-H$-OC-HN R
H.4
=f: HA
d
H; N-H FOC-tin R
i
Ill
Hx
HE3
Fig. 1. Rotamer CH,--CH-_(CH,
notation
for tyrosine
residue in Ala-Tyr
(R = CH,)
and D-Leu-Tyr
(R =
), )-
from those given above. This leads to the errors in the estimated rotamer populations being less than 0.1. Thus use of the described approximations for conformational studies both of metal-free and coordinated dipeptides seems to be quite reasonable and convincing. The conformation behaviour of tyrosine residue in both studieo -stems, i.e. Pd(II)--Ala-Tyr and Pd(II)-D-Leu-Tyr, is very similar to that for the Pd(II)Gly-Tyr system [I]. In 1:l Pd(II)-dipeptide complexes with tridentate Iigand (pH 3-10) the rotamer 3 very considerably increases its population at the expense of 1.A coordinated metal ion stabilizes the gauche conformation of a tyrosine residue in ah cases to almost the same extent (Tables 1 and 2, Fig. 20). The distributions of rotamer populations in all three complexes are equal to each other within the experimental error, i.e.p, = 0.02 -+0.02,~~ = 0.26 -+0.02 and p3 = 0.71 + 0.02. Thus the bulky side-chain groups of a C-termi& amino acid hardly seem to affect tyrosine conformation at all. It is interesting to note that for metal-free D-Leu-Tyr the dipeptide molecule has quite a compact structure. The methyl groups of leucine residue are near the face of the tyrosine ring and this influences the tyrosine side-chain conformation considerably [ 10 ] . In 1:1 complexes with a non-coordinated caxboxyl group (high pH region), the tyrosine residue conformation changes drastically. The rotamer 1 becomes
78
Fig. 2. The most stable conformers for 1:l Pd(II)-dipeptide and bidentate (b) ligand.
complex for tridentate (a)
the most stable (Fig. 2b), at least in Ala-Tyr complex, and its population is much higher than that in metal-free dipeptide, mainly at the expense of rotamer 3 (Table 1). An analogous situation could be in the case of the D-LeuTyr complex, although the data obtained do not permit acceptance of this conclusion without doubts. The chemical shift differences between the 1:l complexes with tidentate and bidentate ligands are most probably caused by the chelate ring conformation changes after hydrolysis of the metal--carboxyl bond, as was found for other similar systems [l, 21. For a metal complex with a tridentate ligand the chelate rings are close to planarity as are those in the Cu(II)-Leu-Tyr complex [5] as well as other systems [3,4, 61. The breaking of the metal-carboxyl bonds can lead to greater deviation from planarity of the remaining chelate rings formed by the C-terminal amino acid. In the 1: 2 Pd(II)-Ala-Tyr complexes the conformation of the tyrosine residue is only slightly different, favouring rotamer 3 at the expense of rotamer 2, compared to the 1: 1 complex at high pH (above 13). There are, however, quite large chemical shift variations mainly for A (0.274 ppm) and X, (rCH (0.183 ppm) protons of tyrosine residue. This suggests that the coordination of the second dipeptide molecule may induce some further conformational changes. The mutual interaction between two dipeptide molecules would lead to some changes in dihedral angle @ characterizing the N-C, bond in the peptide that would put the side-chain tyrosine groups further away from the complex plane. The differences in chemical shifts for CH3 and (rC!H alanine protons in 1 :l and 1: 2 complexes would suggest a decrease in the chelate ring distortion from planarity in the 1: 2 complex as compared to the
79
1: 1 complex with a bidentate ligand or the faster exchange between chelate ring conformers that averages the PMR parameters to values cIose to those for the planar conformer. The crCH proton chemical shift becomes very close to that observed in the 1:l compiex at low pH, i.e. with tridentate ligand (Table 1; see above). In the case of Pd(II)-D-Leu-Tyr complexes the differences in the PMR spectra of high pH 1:l and 1: 2 complexes are more considerable than those for the Pd(II)-AIa-Tyr system. The PMR spectrum of a 1: 2 solution abow pH 13 is of ABX type with distribution of tyrosine conformers similar to that in the 1: 2 Pd(II)-Ala-Tyr complex (Tables 1 and 2). The chemical shift differences, especially for tyrosine X, (uCH (0.431 ppm) and A (0.362 ppm) protons are much greater than those with Ala-Tyr and are caused by the same factors as have been outiined for the complex with the latter ligand. A comparison of these results with those for Pd(II)--Gly-Tyr [l] and Pd(II)--Gly-Phe [2 ] , where almost no differences between “high pH” 1:l and 1: 2 complexes have been observed, has allowed us to conclude that, in mutual interactions between two ligand molecules coordinated to one metal ion, the main role is played by the side chains of C-terminal amino acids. This also explains why in the D-Leu-Tyr case the interactions seem to be more effective. It could be concluded that, in complexes with Pd(II) ion, the bulky sidechain groups will not play any important role in inducing the aromatic ring containing side-chain conformation around the C,-C, bond. It could, however, affect the N-C, rotation via the intramolecular interactions in 1: 2 complexes. Temperature
dependence
of the PMR spectra for Pd(ll)-Gly-Tyr
complexes
The temperature increase for a 1:l solution of Pd(II)--Gly-Tyr (PI-I = 4.2) for which the best spectra could be obtained causes a small reduction in the rotamer 3 population as well as some downfield chemical shifts of (YCH and PCH, tyrosine protons (Table 3). There is almost no change in the chemical shifts associated with CH* glycine protons. Using the Cavanaugh approach [173, the temperature dependence of the rotamer energy differences could be explained by some inter- or intramoIecu1a.r interactions, although their character would not be well-defined [lo, 17, 131. The metal--aromatic ring interaction could be only a small part of the whole set of interactions which might include solute-solvent or the solute-solute ones. The lack of chemical shift temperature variations of the glycine methylene protons is in agreement with earlier assumptions that the chelate ring formed by this ammo-acid residue is almost planar [ 11. All discussed systems are now under CD and ORD studies and the results will be published elsewhere.
Temperature dependence of the PMR parameters, rotamer differences for Pd(II)-Gly-Tyr complex at pH 4.2 "X
iJAB
JAX
JBX
c%+lY
PI
populations
Pa
and rotamer
p3
E,
-E3
energy
E
-E
(kcalmol-')&al~ol-'1 293
171.7
209.5
305.3
14.1
3.0
5.3
230.5
322
172.4
210.1
307.3
14.1
3.1
5.6
230.6
341
173.2
210.7
308.6
14.1
3.3
5.7
230.6
363
174.1
210.5
310.2
14.0
3.3
5.7
230.3
0.04 (0.00) 0.04 (0.00) 0.06 (0.02) 0.06
0.24 CO.081 0.27 (0.13) 0.28 (0.15) 0.28
371
174.5
211.4
311.6
14.3
3.6
5.8
230.3
';.",,"I (0:05) ';A;' (0:18)
0.72 (0.92) 0.69 (0.87) 0.66 (0.83) 0.66 (0.83) ("0%)
1.682 (7) 1.822 (?) 1.625 (2.525) 1.730 (2.687) 1.422 (2.015)
0.639 (1.422) 0.600 (1.216) 0.581 (1.159) 0.618 (1.234) 0.500 (1.071)
ACKNOWLEDGEMENTS
The authors wish to express their sincere thanks to Professor B. JeiowskaTrzebiatowska for encouragement in the work performed. The work was supported by the Polish Academy of Sciences (No. MR l-9). REFERENCES 1 H. Kodowski and M. Jeiowska, Chem. Phys. Lett., 47 (1977) 452. 2 H. Kodowski, G. Formicka-Koziowska and B. Jeiowska-Trzebiatowska, Org. Magn. Resonance, 10 (1978) 146. 3 W. A. Franks and D. van der Helm, Acta Crystallogr. Sect. B, 27 (1970) 1299. 4 D. van der Helm and C. E_ Tatsch, Acta Crystallogr. Sect. B, 28 (1972) 2307. 5 D. van der Helm, S. E. Ealick and J. E. Burks, Acta Crystallogr. Sect. B, 31 (1975) 1013. 6 P. A. Mosset and J. J. Bonnet, Acta Crystallogr. Sect. B, 33 (1977) 2807. 7 M. B. Hursthouse, S. A. A. Jayaweera, G. H. W. Milburn and A. Quick, J. Chem. Sot. D, (1971) 207. 8 J. Feeney, J. Magn. Resonance, 21 (1976) 473. 9 K. G. R. Pachler, Spectrochim. Acta, 20 (1964) 581. 10 B. J. Dale and D. W. Jones, 3. Chem. Sot. Perkin Trans., 2 (1971) 91. 11 L. E. Nance, A. F. Schreiner and G. Frye, Bioinorg. Chem., 3 (1974) 135, and references therein. 12 H. KoZtowski and B. Jeiowska-Trzebiatowska, Chem. Phys. Lett., 42 (1976) 246; H. Koztowski, Inorg. Chim. Acta, 24 (1977) 215. 13 E. W. Wilson Jr. and R. B. Martin, Inorg. Chem., 2 (1970) 528. 14 B. Jeiowska-Trzebiatowska, G. Formicka-Kodowska and H. Koztowski, Bull. Acad. Pal. Sci., Ser. Sci. Chim., in press. 15 L. Rasmussen and C. K. JQrgensen, Acta Chem. Stand., 22 (1968) 2313. 16 P. M. Cotrait and J.-P. Bideau, Acta Crystallogr. Sect. B., 30 (1974) 1024. 17 J. R. Cavanaugh, J. Am. Chem. Sot., 90 (1968) 4533; J. R. Cavanaugh, J. Am. Chem. Sot., 93 (1970) 1488. 18 K. D. Bartle, D. W. Jones and R. L’ Arnie, J. Chem. Sot., Perkin Trans. 2, (1972) 650.
73-80 o Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands
PMR CONFORMATIONAL STUDIES OF Pd(I1) COMPLEXES WITH ALATYR AND D-LEU-TYR DIPEPTJDES
HENRYK
KOZ%OWSKI,
Institute of Chemistry, (Received
MALGORZATA
JEZOWSKA and HELENA SZYSZUK
University of Wrocibw, Joliot-Curie 14, 50-383,
Wroctaw (Poland)
13 March 1978)
ABSTRACT
PMR studies of Pd(II) complexes with Ala-Tyr and D-Lcu-Tyr role of metal ion in inducing tridentate
coordination
the dipeptide
of dipeptide
conformation
(pH 3-10)
revealed the essential in solution. In a complex with
the most stable conformer
residue is that with the aromatic ring in gauche position
to carbgxyl
of a
tyrosine
and amide groups. At
is a bidentate ligand (NH,, N-), the tyrosine residue changes its conformation drastically, and the most stable conformer is that with the aromatic ring in trans position to the carboxyl group. It was found that the C-terminal amino acid does not have
high pH a dipeptide
any considerable
influence
on the conformation
of an N-terminal
amino acid in both kinds
complexes with palladium(D). Increasing the temperature up to 370 K has only a minor effect on the conformer population in solution. of
INTRODUCTION Our recent studies on the Pd(II) complexes with dipeptides containing the aromatic ring in an amino-acid side chain [ 1,2] revealed that the metal ion induced some favourable conformation of such a side chain which varied considerably compared to that found in a metal-free ligand. For the studied complexes, i.e. Pd( II) with glycyl-L-tyrosine (Gly-Tyr), glycyl-L-phenylalanine
(Gly-Phe) and L-phenylalanyl-glycine (Phe-Gly), in the most stable conformer the aromatic ring was above the square planar complex plane. To explain the stabilization of such a rotational isomer (gauche) an interaction between aromatic ring and metal ion has been proposed [ 1,2] . X-ray studies of some Cu( II) complexes with tyrosine [ 3-63 and tryptophane [ 71 containing ligands have shown that the interaction between the metal ion and side-chain group was possible but it was not the rule [5] . The molecules which have the interaction between aromatic ring and copper (II) ion are in the same approximate conformation as that in the Cu(I1) complex with L-leucyl-L-tyrosine (no interaction) [5] . Thus, the conformation of the amino-acid side chain would not be a good criterion for determining whether an interaction will occur. In Pd(I1) complexes, however, such interaction happens more easily because there is no apical coordination to the metal ion, which is the case in cupric complexes [3-G].
74 EXPERIMENTAL Glycyl-L-tyrosine ( Gly-Tyr), L-alanyl-L-tyrosine and D-leucyl-L-tyrosine (D-Leu-Tyr) dipeptides were used as obtained from Fluka AG. K2PdC!14was obtained by crystallization of KC1 and PdClz solutions containing HCl. Analysis showed a purity better than 99%. PMR spectra were recorded on a JEOL 100 MHz JNM PS-100 spectrometer using tert-butyl alcohol as an internal standard. All spectra were measured at 25 * 2”C, and those for Pd(II)Gly-Tyr solutions were measured up to 98°C. The pH was adjusted on a MeraElmat N-5122 pH-meter. PMR spectra of tyrosine oCH$CH2 protons have been treated as of ABX type and they were analyzed on a JEC-6 computer. The concentration of dipeptides was 0.15 mole 1-l in all samples. DzO (99.8%) was used as a solvent. RESULTS The PMR parameters for metal-free Ala-Tyr and D-LeU-TYr at chosen pH are listed in Tables 1 and 2. The rotational isomer notation for tyrosine residue is given in Fig. 1. The fractional population of each isomer can be estimated by the Feeney [8] or Pachler [9] approximation (Tables 1 and 2). The data TABLE 1 ‘H NMR parameters and rotamer populations for Ala-Tyr dipeptide and its complexes with Pd(II) ion (the populations obtained by Feeney’s equations are given in parentheses) PH
VA
vx
iJAB
JAX
JBX
WH@
VCH
0.53 175.2
191.3
336.4
13.9
9.0 5.7
24.8
276.8
2.69 173.2
189.2
331.2
14.0
8.5 5.4
24.6
273.8
5.64
164.8
184.8
311.9
14.0
9.0 4.9
24.5
271.3
8.47
162.8
185.8
312.7
14.0
8.9 4.8
3.9
236.8
10.32 158.7
182.0
311.2
14.0
8.9 4.8
-8.9
213.8
13.40 156.9
177.4
309.4
14.0
8.8 4.6
-7.8
214.6
Pd(il)-Ala-TyrI:1 8.60 167.9 197.9
300.2
13.7
2.9
5.7
4.0
230.0
13.80 161.0
166.5
297.3
13.8
11.8
3.9
-22.3
Pd(II)-Ala-TyrI:2 13.59 133.6 177.7
315.6
13.9
11.5
2.9
-16.1
Pl
Pa
0.58 0.28 (0.64) (0.36) 0.54 0.25 (0.59) (0.40) 0.58 0.21 (0.65) (0.27) 0.57 0.20 (0.64) (0.25) 0.57 0.20 (0.64) (0.25) 0.56 0.18 (0.64) (0.22)
P3 0.14 (0.00) 0.21 (0.01) 0.21 (0.08) 0.23 (0.11) 0.23 (0.11) 0.26 (0.14)
0.03 0.28 0.69 (0.00) (0.12) (0.88) 0.04 218.1 0.84 0.12 (0.97) (0.03) (0.00) 228.2
0.81 (0.96)
0.03 (0.04)
0.16 (0.00)
75 TABLE
2
‘H NMR parameters and rotamer populations for D-Leu-Y&r dipeptide and its complexes with Pd(II) ion
PH
“B
“X
IJAB
JAX
4.04 149.5
205.8
332.0
14.3
12.0 4.0 -51.7
254.7
6.08
148.4
204.4
326.4
14.1
12.0 3.8 -52.9
255.4
9.78
149.1
195.3
324.2
14.2
10.9
4.2
-53.3
200.5
11.62
148.3
183.2
315.4
14.1
9.6
4.2
-48.4
203.5
13.50
148.1
181.2
314.0
14.1
9.8
4.2
-47.4
203.9
Pd(Il)--o-Leu-Tyr 1:1 8.20 164.0 208.6
294.9
13.5
2.8
5.4
-30.4
225.5
13.90
280.0
14.4
-33.4
184.8
11.3 3.3
-28.5
200.1
“A
169.4
Pd(ZI)-D-Leu-Tyr
13.76 133.2
JBX
UCH,h”
“CH
Pl
P2
PO
0.02 (0.00) 0.04 (0.00) 0.14 (0.13) 0.14 (0.19) 0.14 (0.18)
0.11 (0.00) 0.22 (0.08) 0.21 (0.07)
0.02 0.25 (0.00) (0.07) 0.84 (0.92)
0.73 (0.93) 0.16 (0.08)
0.79 (0.93)
0.15 (0.00)
0.75 (0.87) 0.64 (0.73) 0.65 (0.75)
1-2
170.3
323.1
13.7
0.06 (0.07)
obtained are in good agreement with those given by Dale and Jones [lo] for both peptides. ln tyrosine residue, over the whole pH range, the most stable rotamer is that with the carboxyl group and aromatic ring truns to each other (1). The gauche isomer (3) is rather unstable in metal-free peptide, though its population in basic solution is quite considerable at least for the Pachler approximation (Tables 1 and 2). Pd(ll)
: dipeptide
1: 1 molar solutions
Addition of Pd(I1) ions to a dipeptide containing solution changes the PMR spectra of the latter dramatically. For 1: 1 solutions in pH range 3-10, the PMR spectra have the parameters given at pH 8.6 for Pd(II)-Ala-Tyr solutions (Table 1) and at pH 8.20 for Pd(II)-&Leu-Tyr ones (Table 2). The chemical shifts as well as the coupling constant variations in comparison to the metalfree ligand are caused by the metal ion coordination to the dipeptide molecule. The possible coordination sites of Pd(II) ion in both peptides are the amine group, the deprotonated nitrogen of a peptide linkage and the carboxyl group. At any pH value between 3 and 10 all these three donor groups are coordinated to the metal ion as has already been found for similar systems [l, 2,11,12, 13) . Also d-d transition energies found for both kinds of solutions, i.e. 332 and 333 nm for Pd(II)--Ala-Tyr and Pd(If)-D-Leu-Tyr, respectively, may suggest the trident&e coordination by NHP, N- and COO- donors [2,11,13].
76
The conformation of a tyrosine residue of those tridentately coordinated dipeptides is quite different to that found in metal-free ligands (Tables 1 and 2). There is a considerable increase of the rotamer 3 (gauche) population, and the rotamer 1, most stable in the metal-free ligands, is almost absent in the complex solution (p I <0.05). An increase of pH above 13 in 1:l Pd( II)-Ala-Tyr solutions leads to the hydrolysis of metal-car-boxy1 bonds [ 1, 2, 11, 12,133 , which results in quite drastic changes in the PMR spectrum (Table 2). There is an upfield chemical shift of all protons of dipeptide molecules except aCH-Tyr (Table 1). There is also a large increase in JAx value (2.9 to 11.8) and, as a consequence, a large increase of rotamer 1 population (-O-t- 0.9). The rotamer 2, which was quite appreciably populated for tridentately coordinated and metal-free Ala-Tyr, vanished almost completely (pl = 0.02; Table 1). Similar variations in the PMR spectrum were found for the 1 :l Pd( 11)-DLeu-Tyr solution above pH 13. The spectrum of the tyrosine residue, however, is of A2X type (or rather deceptively simple ABX) and it is not possible to distinguish the contributions of rotamers 1 and 2. In both cases, the decrease in gauche rotamer population was either to the same or to an even lower value than that was found for the metal-free ligand in the same pH region (Tables 1 and 2). Pd(II)
: dipeptide
1:2 molar solutions
The PMR spectra of 1:2 metal to peptide molar solutions up to pH 12 consist of two sets of lines, one for the 1:l complex described above and the other for metal-free ligand. Pd(II)-peptide systems are quite inert on an NMR time-scale and we usually observe a separate PMR spectrum for each chemical species in solution. Above pH I.3 only one single spectrum is observed, suggesting the coordination of two dipeptide molecules to a metal ion. A break of the metalcarboxyl bond makes the formation of two chelates with two ligand molecules coordinated by NH2 and N- possible [l, 2,121. Coordination of four nitrogen donors to Pd(I1) ion is also supported by the absorption spectra. The d-d transition energies (h <300 nm) found for 1:2 Pd(II):Ala-Tyr and Pd(II):D-Leu-Tyr are close to the values found for Pd(II) ion with four coordinated nitrogens [Z, 13,14,15]. An overlapping of the d-d transition band with that for WV* in the aromatic ring (Tyr) does not permit the precise calculation of the former transition energy. DISCUSSfON X-ray studies have shown that the metal ions coordinated to a peptide molecule do not affect considerably the dihedral angles in tyrosine residue [3-6,16] , which were assumed to be 60”, 180” and 300” (Fig. l), and that in coordinated or metal-free ligands these angles could deviate by about 3-8”
77
CO’JH I
W-kOH COOH &HL OH
H0 d H;N-H$-OC-HN R
H.4
=f: HA
d
H; N-H FOC-tin R
i
Ill
Hx
HE3
Fig. 1. Rotamer CH,--CH-_(CH,
notation
for tyrosine
residue in Ala-Tyr
(R = CH,)
and D-Leu-Tyr
(R =
), )-
from those given above. This leads to the errors in the estimated rotamer populations being less than 0.1. Thus use of the described approximations for conformational studies both of metal-free and coordinated dipeptides seems to be quite reasonable and convincing. The conformation behaviour of tyrosine residue in both studieo -stems, i.e. Pd(II)--Ala-Tyr and Pd(II)-D-Leu-Tyr, is very similar to that for the Pd(II)Gly-Tyr system [I]. In 1:l Pd(II)-dipeptide complexes with tridentate Iigand (pH 3-10) the rotamer 3 very considerably increases its population at the expense of 1.A coordinated metal ion stabilizes the gauche conformation of a tyrosine residue in ah cases to almost the same extent (Tables 1 and 2, Fig. 20). The distributions of rotamer populations in all three complexes are equal to each other within the experimental error, i.e.p, = 0.02 -+0.02,~~ = 0.26 -+0.02 and p3 = 0.71 + 0.02. Thus the bulky side-chain groups of a C-termi& amino acid hardly seem to affect tyrosine conformation at all. It is interesting to note that for metal-free D-Leu-Tyr the dipeptide molecule has quite a compact structure. The methyl groups of leucine residue are near the face of the tyrosine ring and this influences the tyrosine side-chain conformation considerably [ 10 ] . In 1:1 complexes with a non-coordinated caxboxyl group (high pH region), the tyrosine residue conformation changes drastically. The rotamer 1 becomes
78
Fig. 2. The most stable conformers for 1:l Pd(II)-dipeptide and bidentate (b) ligand.
complex for tridentate (a)
the most stable (Fig. 2b), at least in Ala-Tyr complex, and its population is much higher than that in metal-free dipeptide, mainly at the expense of rotamer 3 (Table 1). An analogous situation could be in the case of the D-LeuTyr complex, although the data obtained do not permit acceptance of this conclusion without doubts. The chemical shift differences between the 1:l complexes with tidentate and bidentate ligands are most probably caused by the chelate ring conformation changes after hydrolysis of the metal--carboxyl bond, as was found for other similar systems [l, 21. For a metal complex with a tridentate ligand the chelate rings are close to planarity as are those in the Cu(II)-Leu-Tyr complex [5] as well as other systems [3,4, 61. The breaking of the metal-carboxyl bonds can lead to greater deviation from planarity of the remaining chelate rings formed by the C-terminal amino acid. In the 1: 2 Pd(II)-Ala-Tyr complexes the conformation of the tyrosine residue is only slightly different, favouring rotamer 3 at the expense of rotamer 2, compared to the 1: 1 complex at high pH (above 13). There are, however, quite large chemical shift variations mainly for A (0.274 ppm) and X, (rCH (0.183 ppm) protons of tyrosine residue. This suggests that the coordination of the second dipeptide molecule may induce some further conformational changes. The mutual interaction between two dipeptide molecules would lead to some changes in dihedral angle @ characterizing the N-C, bond in the peptide that would put the side-chain tyrosine groups further away from the complex plane. The differences in chemical shifts for CH3 and (rC!H alanine protons in 1 :l and 1: 2 complexes would suggest a decrease in the chelate ring distortion from planarity in the 1: 2 complex as compared to the
79
1: 1 complex with a bidentate ligand or the faster exchange between chelate ring conformers that averages the PMR parameters to values cIose to those for the planar conformer. The crCH proton chemical shift becomes very close to that observed in the 1:l compiex at low pH, i.e. with tridentate ligand (Table 1; see above). In the case of Pd(II)-D-Leu-Tyr complexes the differences in the PMR spectra of high pH 1:l and 1: 2 complexes are more considerable than those for the Pd(II)-AIa-Tyr system. The PMR spectrum of a 1: 2 solution abow pH 13 is of ABX type with distribution of tyrosine conformers similar to that in the 1: 2 Pd(II)-Ala-Tyr complex (Tables 1 and 2). The chemical shift differences, especially for tyrosine X, (uCH (0.431 ppm) and A (0.362 ppm) protons are much greater than those with Ala-Tyr and are caused by the same factors as have been outiined for the complex with the latter ligand. A comparison of these results with those for Pd(II)--Gly-Tyr [l] and Pd(II)--Gly-Phe [2 ] , where almost no differences between “high pH” 1:l and 1: 2 complexes have been observed, has allowed us to conclude that, in mutual interactions between two ligand molecules coordinated to one metal ion, the main role is played by the side chains of C-terminal amino acids. This also explains why in the D-Leu-Tyr case the interactions seem to be more effective. It could be concluded that, in complexes with Pd(II) ion, the bulky sidechain groups will not play any important role in inducing the aromatic ring containing side-chain conformation around the C,-C, bond. It could, however, affect the N-C, rotation via the intramolecular interactions in 1: 2 complexes. Temperature
dependence
of the PMR spectra for Pd(ll)-Gly-Tyr
complexes
The temperature increase for a 1:l solution of Pd(II)--Gly-Tyr (PI-I = 4.2) for which the best spectra could be obtained causes a small reduction in the rotamer 3 population as well as some downfield chemical shifts of (YCH and PCH, tyrosine protons (Table 3). There is almost no change in the chemical shifts associated with CH* glycine protons. Using the Cavanaugh approach [173, the temperature dependence of the rotamer energy differences could be explained by some inter- or intramoIecu1a.r interactions, although their character would not be well-defined [lo, 17, 131. The metal--aromatic ring interaction could be only a small part of the whole set of interactions which might include solute-solvent or the solute-solute ones. The lack of chemical shift temperature variations of the glycine methylene protons is in agreement with earlier assumptions that the chelate ring formed by this ammo-acid residue is almost planar [ 11. All discussed systems are now under CD and ORD studies and the results will be published elsewhere.
Temperature dependence of the PMR parameters, rotamer differences for Pd(II)-Gly-Tyr complex at pH 4.2 "X
iJAB
JAX
JBX
c%+lY
PI
populations
Pa
and rotamer
p3
E,
-E3
energy
E
-E
(kcalmol-')&al~ol-'1 293
171.7
209.5
305.3
14.1
3.0
5.3
230.5
322
172.4
210.1
307.3
14.1
3.1
5.6
230.6
341
173.2
210.7
308.6
14.1
3.3
5.7
230.6
363
174.1
210.5
310.2
14.0
3.3
5.7
230.3
0.04 (0.00) 0.04 (0.00) 0.06 (0.02) 0.06
0.24 CO.081 0.27 (0.13) 0.28 (0.15) 0.28
371
174.5
211.4
311.6
14.3
3.6
5.8
230.3
';.",,"I (0:05) ';A;' (0:18)
0.72 (0.92) 0.69 (0.87) 0.66 (0.83) 0.66 (0.83) ("0%)
1.682 (7) 1.822 (?) 1.625 (2.525) 1.730 (2.687) 1.422 (2.015)
0.639 (1.422) 0.600 (1.216) 0.581 (1.159) 0.618 (1.234) 0.500 (1.071)
ACKNOWLEDGEMENTS
The authors wish to express their sincere thanks to Professor B. JeiowskaTrzebiatowska for encouragement in the work performed. The work was supported by the Polish Academy of Sciences (No. MR l-9). REFERENCES 1 H. Kodowski and M. Jeiowska, Chem. Phys. Lett., 47 (1977) 452. 2 H. Kodowski, G. Formicka-Koziowska and B. Jeiowska-Trzebiatowska, Org. Magn. Resonance, 10 (1978) 146. 3 W. A. Franks and D. van der Helm, Acta Crystallogr. Sect. B, 27 (1970) 1299. 4 D. van der Helm and C. E_ Tatsch, Acta Crystallogr. Sect. B, 28 (1972) 2307. 5 D. van der Helm, S. E. Ealick and J. E. Burks, Acta Crystallogr. Sect. B, 31 (1975) 1013. 6 P. A. Mosset and J. J. Bonnet, Acta Crystallogr. Sect. B, 33 (1977) 2807. 7 M. B. Hursthouse, S. A. A. Jayaweera, G. H. W. Milburn and A. Quick, J. Chem. Sot. D, (1971) 207. 8 J. Feeney, J. Magn. Resonance, 21 (1976) 473. 9 K. G. R. Pachler, Spectrochim. Acta, 20 (1964) 581. 10 B. J. Dale and D. W. Jones, 3. Chem. Sot. Perkin Trans., 2 (1971) 91. 11 L. E. Nance, A. F. Schreiner and G. Frye, Bioinorg. Chem., 3 (1974) 135, and references therein. 12 H. KoZtowski and B. Jeiowska-Trzebiatowska, Chem. Phys. Lett., 42 (1976) 246; H. Koztowski, Inorg. Chim. Acta, 24 (1977) 215. 13 E. W. Wilson Jr. and R. B. Martin, Inorg. Chem., 2 (1970) 528. 14 B. Jeiowska-Trzebiatowska, G. Formicka-Kodowska and H. Koztowski, Bull. Acad. Pal. Sci., Ser. Sci. Chim., in press. 15 L. Rasmussen and C. K. JQrgensen, Acta Chem. Stand., 22 (1968) 2313. 16 P. M. Cotrait and J.-P. Bideau, Acta Crystallogr. Sect. B., 30 (1974) 1024. 17 J. R. Cavanaugh, J. Am. Chem. Sot., 90 (1968) 4533; J. R. Cavanaugh, J. Am. Chem. Sot., 93 (1970) 1488. 18 K. D. Bartle, D. W. Jones and R. L’ Arnie, J. Chem. Sot., Perkin Trans. 2, (1972) 650.