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A solid-state 205TI NMR study of the Tl+-lasalocid− and Tl+-gramicidin complexes
JOURNAL
OF MAGNETIC
RESONANCE
64, 120- 123 ( 1985)
A Solid-State *“Tl NMR Study of the TV-Lasalocidand Tl+-Gramicidin Complexes J.F. HINTON,K.R.METZ,G.
L.TuRNER,D.L.
BENNETT, AND F.S.MILLETT
Department of Chemistry, University of Arkansas. Fayetteville, Arkansas 72701 Received December 26, 1984
W e have recently shown that 205Tl+ NMR spectroscopy is a good technique for studying the interaction between antibiotic ionophores and metal cations (1-5). It was found that the resonance frequency of the Tl+ ion, complexed by an antibiotic, is directly related to the basicity of the functional groups used by the antibiotic to bind the cation. 205T1+NMR spectroscopy has been used to determine the number of binding sites and the thermodynamics for the binding process for the complex formed between the antibiotic gramicidin in solution and in model membranes and the Tl+ ion (4, 5). To more fully understand the relaxation characteristics (T,) of the 205T1+ nucleus in antibiotic complexes and to obtain information about the geometry of the binding site it has become necessary to obtain and interpret the solid-state 205T1+NMR spectra of some antibiotic complexes. W e report the results of a solid-state 205Tlf NMR study of two antibiotic complexes, prepared with lasalocid and gramicidin-A, that represent the extremes in structure and function. In membranes, antibiotics may function as channel formers or ion carriers to facilitate ion transport across the membrane. Lasalocid (X-537A) (Fig. 1) is a carboxylic ionophore that acts as an ion carrier and has been used in the study of transport processes in physiological systems (6-12) while gramicidin-A is a well known channel former (12). Not only are these two antibiotics functionally different, they are also structurally different. Gramicidin-A (Fig. 1) is a neutral polypeptide which forms helical dimers that act as ion channels in membrane systems. The interior of the channel is lined with carbonyl groups which bind electrostatically with the cation. The tight binding site is located at the channel entrance (3, 13-1.5). No crystal-structure data exist for the Tl+-gramicidin-A complex to provide the binding site geometry. X-ray crystallographic studies of K+ and Cs+ gramidicin-A complexes are not of sufficiently high resolution to determine the symmetry of the binding site (16). No X-ray crystallographic study has been performed with the Tl+-lasalocidcomplex; however, proton NMR studies of the complex (1, 17-19) and analogies drawn from X-ray studies of the sodium salt (20) suggest that the Tl’ ion is complexed by two lasalocid anions forming a binding site consistent with nonaxial symmetry. Spin-lattice relaxation time measurements of 205T1+ in the complex indicate that relaxation is dominated by the chemical shift anisotropy mechanism 0022-2364185 $3.00 Copyright 0 1985 by Academic Press, Inc. All rights of reproduction in any form reserved.
120
121
NOTES
(A) HO
OH
% “s
“f
9"3'2 y
(YH,), (yH312 (FH3 I2
(FH3'2 Ii
FH3
FH
FH3
fli
CH
$tl
HCONHCH$NH~H$NHCH$NHCH$NHCH$NHCH$NHbH$NHCH 0
TRY
0
0
TRY
0
0
TRY
0
0
TRY
H
FIG. 1. Antibiotic structural formulas. (A) Lasalocid-; (B) gramicidin-A.
(1). Consequently, it is of interest to determine how large the chemical shift anisotropy is in this Tl+ antibiotic salt and whether or not nonaxial symmetry is suggested by the solid-state 205TI+powder pattern.
FIG.
2. Solid-state zo5Tl powder pattern of the TV-lasalocid- complex.
122
NOTES TABLE 033
a22
Complex
W/ppmY
Wz/ppm)
LasalocidGramicidin
577298471806.3 577274181764.2
576941831188.1 576996641283.1
1 011
Whpm) 576669 121284.7 57689672/109.0
%o
W/mm) 5769698 11236.6 577055851385.7
A0 (Hzhpm) 443691854.6 294741567.7
’ Reference is Tl+ in H20 at infinite dilution.
The details of the preparation of the Tl+-lasalocid- and Tl+-gramicidin-A complexes are described in the literature (I, J-5). A modified Bruker HFX-90 spectrometer (2, 21) was used to obtain the solid-state spectra. A computer program, POWDER, written in this laboratory was used to obtain the principal elements of the chemical-shift tensor (a, ,, u22, and u&. Figure 2 shows the solid-state powder pattern for 205Tl+ in the Tl+-lasalocidcomplex. The powder pattern is of the nonaxially symmetric type (i.e., ull > u22 > (~33).From the data obtained by computer fit of the powder pattern (Table I), the chemical shift anisotropy, Au = u33 - l/2( uII + (Tag),is 854.6 ppm. A value of Au this large would certainly make the chemical-shift-anisotropy mechanism an important contributor to the overall relaxation of the 205T1+nucleus in the complex in solution. The isotropic chemical shift, uiso = 1/3(urr + (~22+ u33), is 236.6 ppm compared to a value of 297.6 ppm for the complex in CHC13 solution. There is obviously a significant structural change in the complex in going from the solution to the solid state. This appears to be typical at least for some thallium salts; for example, a chemical shift change of about 400 ppm occurs for TlC13 in going from an aqueous solution to the solid state (22). The experimental powder pattern for the Tl+-gramicidin-A complex, shown in Fig. 3, could be fit only by assuming nonaxial symmetry about the binding site.
FIG. 3. Solid-state “‘Tl
powder pattern of the Tl+-gramicidin-A
complex.
NOTES
123
The chemical shift anisotropy, Au, for this complex is 567.7 ppm (see Table 1 for pertinent data). One would expect to observe a powder pattern of axial symmetry only if the Tl+ ion were located on the channel axis surrounded by symmetrically positioned binding groups with the anion also located on the channel axis. This does not seem to be the case with the solid complex. ACKNOWLEDGMENT We acknowledge the National
Science Foundation
for support through Grant PCM-8300065.
REFERENCES 1. 2. 3. 4. 5. 6. 7. 8. 9.
10. II. 12. 13. 14. 15. 16. 17. 18.
19. 20. 21. 22.
R. W. BRIGGS, F. A. ETZKORN, AND J. F. HINTON, J. Magn. Reson. 37, 523 (1980). J. F. HINTON, K. R. METZ, AND F. S. MILLETT, J. Magn. Reson. 44, 2 17 (198 1). J. F. HINTON, G. L. TURNER, AND F. S. MILLETT, J. Magn. Reson. 45, 42 (1981). G. L. TURNER, J. F. HINTON, AND F. S. MILLETT, Biochemistry 21, 646 (1982). J. F. HINTON, G. YOUNG, AND F. S. MILLETT, Biochemistry 21, 651 (1982). A. H. CASSWELL AND B. C. PRESSMAN, B&hem. Biophys. Res. Commun. 49,292 (1972). A. SCARPA AND G. INESI, FEBS Lett. 22, 273 (1972). M. L. ENTMAN, P. C. GILLETTE, E. T. WALLICK, B. C. PRESSMAN, AND A. SCHWARTZ, Biochem. Biophys. Res. Commun. 48, 847 (1972). H. KITA AND W. VAN DER KL~~T, Nature (London) 250,658 (1974). B. C. PRESSMAN, Ann. N.Y. Acad. Sci. 147, 829 (1969). N. T. DE GUZMAN AND B. C. PRESSMAN, Circulation 46, 1072 (1974). A. FINKELSTEIN AND 0. S. ANDERSEN, J. Membr. Biol. 59, 155 (1981). 0. S. ANDERSEN, E. W. BARRETT, AND L. B. WEISS, Biophys. J. 33, 63 (1981). D. W. URRY, K. V. PRASAD, AND T. L. TRAPANE, Proc. Natl. Acad. Sci. USA 79, 390 (1982). D. W. URRY, J. T. WALKER, AND T. L. TRAPANE, J. Membr. Biol. 69, 225 (1982). R. E. KOEPPE, J. M. BERG, K. 0. HODGSON, AND L. STRYER, Nature (London) 279, 723 (1979). D. J. PATEL AND C. SHEN, Proc. Natl. Acad. Sci. USA 73, 1786 (1976). C. SHEN AND D. J. PATEL, Proc. Natl. Acad. Sci. USA 73, 4277 (1976). J. Y. LALLEMAND AND V. MICHON, J. Chem. Res. (M), 208 1 (1978); J. Chem. Res. (S). 162 (1978). G. D. SMITH, W. L. DUAX AND S. FORTIER, J. Am. Chem. Sot. 100, 6725 (1978). K. R. METZ AND J. F. HINTON, J. Magn. Reson. 45, 229 (1981). J. GLASER AND U. HENRIKSSON, J. Am. Chem. Sot. 103,6642 (1981).
OF MAGNETIC
RESONANCE
64, 120- 123 ( 1985)
A Solid-State *“Tl NMR Study of the TV-Lasalocidand Tl+-Gramicidin Complexes J.F. HINTON,K.R.METZ,G.
L.TuRNER,D.L.
BENNETT, AND F.S.MILLETT
Department of Chemistry, University of Arkansas. Fayetteville, Arkansas 72701 Received December 26, 1984
W e have recently shown that 205Tl+ NMR spectroscopy is a good technique for studying the interaction between antibiotic ionophores and metal cations (1-5). It was found that the resonance frequency of the Tl+ ion, complexed by an antibiotic, is directly related to the basicity of the functional groups used by the antibiotic to bind the cation. 205T1+NMR spectroscopy has been used to determine the number of binding sites and the thermodynamics for the binding process for the complex formed between the antibiotic gramicidin in solution and in model membranes and the Tl+ ion (4, 5). To more fully understand the relaxation characteristics (T,) of the 205T1+ nucleus in antibiotic complexes and to obtain information about the geometry of the binding site it has become necessary to obtain and interpret the solid-state 205T1+NMR spectra of some antibiotic complexes. W e report the results of a solid-state 205Tlf NMR study of two antibiotic complexes, prepared with lasalocid and gramicidin-A, that represent the extremes in structure and function. In membranes, antibiotics may function as channel formers or ion carriers to facilitate ion transport across the membrane. Lasalocid (X-537A) (Fig. 1) is a carboxylic ionophore that acts as an ion carrier and has been used in the study of transport processes in physiological systems (6-12) while gramicidin-A is a well known channel former (12). Not only are these two antibiotics functionally different, they are also structurally different. Gramicidin-A (Fig. 1) is a neutral polypeptide which forms helical dimers that act as ion channels in membrane systems. The interior of the channel is lined with carbonyl groups which bind electrostatically with the cation. The tight binding site is located at the channel entrance (3, 13-1.5). No crystal-structure data exist for the Tl+-gramicidin-A complex to provide the binding site geometry. X-ray crystallographic studies of K+ and Cs+ gramidicin-A complexes are not of sufficiently high resolution to determine the symmetry of the binding site (16). No X-ray crystallographic study has been performed with the Tl+-lasalocidcomplex; however, proton NMR studies of the complex (1, 17-19) and analogies drawn from X-ray studies of the sodium salt (20) suggest that the Tl’ ion is complexed by two lasalocid anions forming a binding site consistent with nonaxial symmetry. Spin-lattice relaxation time measurements of 205T1+ in the complex indicate that relaxation is dominated by the chemical shift anisotropy mechanism 0022-2364185 $3.00 Copyright 0 1985 by Academic Press, Inc. All rights of reproduction in any form reserved.
120
121
NOTES
(A) HO
OH
% “s
“f
9"3'2 y
(YH,), (yH312 (FH3 I2
(FH3'2 Ii
FH3
FH
FH3
fli
CH
$tl
HCONHCH$NH~H$NHCH$NHCH$NHCH$NHCH$NHbH$NHCH 0
TRY
0
0
TRY
0
0
TRY
0
0
TRY
H
FIG. 1. Antibiotic structural formulas. (A) Lasalocid-; (B) gramicidin-A.
(1). Consequently, it is of interest to determine how large the chemical shift anisotropy is in this Tl+ antibiotic salt and whether or not nonaxial symmetry is suggested by the solid-state 205TI+powder pattern.
FIG.
2. Solid-state zo5Tl powder pattern of the TV-lasalocid- complex.
122
NOTES TABLE 033
a22
Complex
W/ppmY
Wz/ppm)
LasalocidGramicidin
577298471806.3 577274181764.2
576941831188.1 576996641283.1
1 011
Whpm) 576669 121284.7 57689672/109.0
%o
W/mm) 5769698 11236.6 577055851385.7
A0 (Hzhpm) 443691854.6 294741567.7
’ Reference is Tl+ in H20 at infinite dilution.
The details of the preparation of the Tl+-lasalocid- and Tl+-gramicidin-A complexes are described in the literature (I, J-5). A modified Bruker HFX-90 spectrometer (2, 21) was used to obtain the solid-state spectra. A computer program, POWDER, written in this laboratory was used to obtain the principal elements of the chemical-shift tensor (a, ,, u22, and u&. Figure 2 shows the solid-state powder pattern for 205Tl+ in the Tl+-lasalocidcomplex. The powder pattern is of the nonaxially symmetric type (i.e., ull > u22 > (~33).From the data obtained by computer fit of the powder pattern (Table I), the chemical shift anisotropy, Au = u33 - l/2( uII + (Tag),is 854.6 ppm. A value of Au this large would certainly make the chemical-shift-anisotropy mechanism an important contributor to the overall relaxation of the 205T1+nucleus in the complex in solution. The isotropic chemical shift, uiso = 1/3(urr + (~22+ u33), is 236.6 ppm compared to a value of 297.6 ppm for the complex in CHC13 solution. There is obviously a significant structural change in the complex in going from the solution to the solid state. This appears to be typical at least for some thallium salts; for example, a chemical shift change of about 400 ppm occurs for TlC13 in going from an aqueous solution to the solid state (22). The experimental powder pattern for the Tl+-gramicidin-A complex, shown in Fig. 3, could be fit only by assuming nonaxial symmetry about the binding site.
FIG. 3. Solid-state “‘Tl
powder pattern of the Tl+-gramicidin-A
complex.
NOTES
123
The chemical shift anisotropy, Au, for this complex is 567.7 ppm (see Table 1 for pertinent data). One would expect to observe a powder pattern of axial symmetry only if the Tl+ ion were located on the channel axis surrounded by symmetrically positioned binding groups with the anion also located on the channel axis. This does not seem to be the case with the solid complex. ACKNOWLEDGMENT We acknowledge the National
Science Foundation
for support through Grant PCM-8300065.
REFERENCES 1. 2. 3. 4. 5. 6. 7. 8. 9.
10. II. 12. 13. 14. 15. 16. 17. 18.
19. 20. 21. 22.
R. W. BRIGGS, F. A. ETZKORN, AND J. F. HINTON, J. Magn. Reson. 37, 523 (1980). J. F. HINTON, K. R. METZ, AND F. S. MILLETT, J. Magn. Reson. 44, 2 17 (198 1). J. F. HINTON, G. L. TURNER, AND F. S. MILLETT, J. Magn. Reson. 45, 42 (1981). G. L. TURNER, J. F. HINTON, AND F. S. MILLETT, Biochemistry 21, 646 (1982). J. F. HINTON, G. YOUNG, AND F. S. MILLETT, Biochemistry 21, 651 (1982). A. H. CASSWELL AND B. C. PRESSMAN, B&hem. Biophys. Res. Commun. 49,292 (1972). A. SCARPA AND G. INESI, FEBS Lett. 22, 273 (1972). M. L. ENTMAN, P. C. GILLETTE, E. T. WALLICK, B. C. PRESSMAN, AND A. SCHWARTZ, Biochem. Biophys. Res. Commun. 48, 847 (1972). H. KITA AND W. VAN DER KL~~T, Nature (London) 250,658 (1974). B. C. PRESSMAN, Ann. N.Y. Acad. Sci. 147, 829 (1969). N. T. DE GUZMAN AND B. C. PRESSMAN, Circulation 46, 1072 (1974). A. FINKELSTEIN AND 0. S. ANDERSEN, J. Membr. Biol. 59, 155 (1981). 0. S. ANDERSEN, E. W. BARRETT, AND L. B. WEISS, Biophys. J. 33, 63 (1981). D. W. URRY, K. V. PRASAD, AND T. L. TRAPANE, Proc. Natl. Acad. Sci. USA 79, 390 (1982). D. W. URRY, J. T. WALKER, AND T. L. TRAPANE, J. Membr. Biol. 69, 225 (1982). R. E. KOEPPE, J. M. BERG, K. 0. HODGSON, AND L. STRYER, Nature (London) 279, 723 (1979). D. J. PATEL AND C. SHEN, Proc. Natl. Acad. Sci. USA 73, 1786 (1976). C. SHEN AND D. J. PATEL, Proc. Natl. Acad. Sci. USA 73, 4277 (1976). J. Y. LALLEMAND AND V. MICHON, J. Chem. Res. (M), 208 1 (1978); J. Chem. Res. (S). 162 (1978). G. D. SMITH, W. L. DUAX AND S. FORTIER, J. Am. Chem. Sot. 100, 6725 (1978). K. R. METZ AND J. F. HINTON, J. Magn. Reson. 45, 229 (1981). J. GLASER AND U. HENRIKSSON, J. Am. Chem. Sot. 103,6642 (1981).