The Role of Cation- Interactions in Biomolecular Association. Design of Peptides Favoring Interactions between Cationic and Aromatic Am
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- 作者：Ekaterina V. Pletneva ; Alain T. Laederach ; D. Bruce Fulton ; and Nenad M. Kosti
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- 刊名：Journal of the American Chemical Society
- 出版年：2001
- 出版时间：July 4, 2001
- 年：2001
- 卷：123
- 期：26
- 页码：6232 - 6245
- 全文大小：362K
- 年卷期：v.123,no.26(July 4, 2001)
- ISSN：1520-5126
文摘
Cation-
interactions between amino acid side chains are increasingly being recognized as importantstructural and functional features of proteins and other biomolecules. Although these interactions have beenfound in static protein structures, they have not yet been detected in dynamic biomolecular systems. Wedetermined, by 1H NMR spectroscopic titrations, the energies of cation- interactions of the amino acidderivative AcLysOMe (1) with AcPheOEt (2) and with AcTyrOEt (3) in aqueous and three organic solvents.The interaction energy is substantial; it ranges from -2.1 to -3.4 kcal/mol and depends only slightly on thedielectric constant of the solvent. To assess the effects of auxiliary interactions and structural preorganizationon formation of cation- interactions, we studied these interactions in the association of pentapeptides. Uponbinding of the positively-charged peptide AcLysLysLysLysLysNH2 (5) to the negatively-charged partnerAcAspAspXAspAspNH2 (6), in which X is Leu (6a), Tyr (6b), and Phe (6c), multiple interactions occur.Association of the two pentapeptides is dynamic. Free peptides and their complex are in fast exchange on theNMR time-scale, and 2D 1H ROESY spectra of the complex of the two pentapeptides do not show intermolecularROESY peaks. Perturbations of the chemical shifts indicated that the aromatic groups in peptides 6b and 6cwere affected by the association with 5. The association constants KA for 5 with 6a and with 6b are nearlyequal, (4.0 ± 0.7) × 103 and (5.0 ± 1.0) × 103 M-1, respectively, while KA for 5 with 6c is larger, (8.3 ± 1.3)× 103 M-1. Molecular-dynamics (MD) simulations of the pentapeptide pairs confirmed that their associationis dynamic and showed that cation- contacts between the two peptides are stereochemically possible. Atransient complex between 5 and 6 with a prominent cation- interaction, obtained from MD simulations,was used as a template to design cyclic peptides CX featuring persistent cation- interactions. The cyclicpeptide CX had a sequence AcLysCys(Lys)3Gly(Asp)2XC ysAspNH2, in which X is Tyr, Phe, and Leu. Thefirst two peptides do, but the third does not, contain the aromatic residue capable of interacting with a cationicLys residue. This covalent construct offered conformational stability over the noncovalent complexes andallowed thorough studies by 2D NMR spectroscopy. Multiple conformations of the cyclic peptides CTyr andCPhe are in slow exchange on the NMR time-scale. In one of these conformations, cation- interaction betweenLys3 and Tyr9/Phe9 is clearly evident. Multiple NOEs between the side chains of residues 3 and 9 are observed;chemical-shift changes are consistent with the placement of the side chain of Lys3 over the aromatic ring. Incontrast, the cyclic peptide CLeu showed no evidence for close approach of the side chains of Lys3 and Leu9.The cation- interaction persists in both DMSO and aqueous solvents. When the disulfide bond in the cyclicpeptide CPhe was removed, the cation- interaction in the acyclic peptide ACPhe remained. To test the reliabilityof the pKa criterion for the existence of cation- interactions, we determined residue-specific pKa values ofall four Lys side chains in all three cyclic peptides CX. While NOE cross-peaks and perturbations of thechemical shifts clearly show the existence of the cation- interaction, pKa values of Lys3 in CTyr and in CPhediffer only marginally from those values of other lysines in these dynamic peptides. Our experimental resultswith dynamic peptide systems highlight the role of cation- interactions in both intermolecular recognitionat the protein-protein interface and intramolecular processes such as protein folding.
interactions between amino acid side chains are increasingly being recognized as importantstructural and functional features of proteins and other biomolecules. Although these interactions have beenfound in static protein structures, they have not yet been detected in dynamic biomolecular systems. Wedetermined, by 1H NMR spectroscopic titrations, the energies of cation- interactions of the amino acidderivative AcLysOMe (1) with AcPheOEt (2) and with AcTyrOEt (3) in aqueous and three organic solvents.The interaction energy is substantial; it ranges from -2.1 to -3.4 kcal/mol and depends only slightly on thedielectric constant of the solvent. To assess the effects of auxiliary interactions and structural preorganizationon formation of cation- interactions, we studied these interactions in the association of pentapeptides. Uponbinding of the positively-charged peptide AcLysLysLysLysLysNH2 (5) to the negatively-charged partnerAcAspAspXAspAspNH2 (6), in which X is Leu (6a), Tyr (6b), and Phe (6c), multiple interactions occur.Association of the two pentapeptides is dynamic. Free peptides and their complex are in fast exchange on theNMR time-scale, and 2D 1H ROESY spectra of the complex of the two pentapeptides do not show intermolecularROESY peaks. Perturbations of the chemical shifts indicated that the aromatic groups in peptides 6b and 6cwere affected by the association with 5. The association constants KA for 5 with 6a and with 6b are nearlyequal, (4.0 ± 0.7) × 103 and (5.0 ± 1.0) × 103 M-1, respectively, while KA for 5 with 6c is larger, (8.3 ± 1.3)× 103 M-1. Molecular-dynamics (MD) simulations of the pentapeptide pairs confirmed that their associationis dynamic and showed that cation- contacts between the two peptides are stereochemically possible. Atransient complex between 5 and 6 with a prominent cation- interaction, obtained from MD simulations,was used as a template to design cyclic peptides CX featuring persistent cation- interactions. The cyclicpeptide CX had a sequence interaction betweenLys3 and Tyr9/Phe9 is clearly evident. Multiple NOEs between the side chains of residues 3 and 9 are observed;chemical-shift changes are consistent with the placement of the side chain of Lys3 over the aromatic ring. Incontrast, the cyclic peptide CLeu showed no evidence for close approach of the side chains of Lys3 and Leu9.The cation- interaction persists in both DMSO and aqueous solvents. When the disulfide bond in the cyclicpeptide CPhe was removed, the cation- interaction in the acyclic peptide ACPhe remained. To test the reliabilityof the pKa criterion for the existence of cation- interactions, we determined residue-specific pKa values ofall four Lys side chains in all three cyclic peptides CX. While NOE cross-peaks and perturbations of thechemical shifts clearly show the existence of the cation- interaction, pKa values of Lys3 in CTyr and in CPhediffer only marginally from those values of other lysines in these dynamic peptides. Our experimental resultswith dynamic peptide systems highlight the role of cation- interactions in both intermolecular recognitionat the protein-protein interface and intramolecular processes such as protein folding.