Controlled Peptide Solvation in Portion-Mixing Libraries of FRET Peptides: Improved Specificity Determination for Dengue 2 Virus NS2B-NS3 Protease and Human Cathepsin S

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• 作者：Fabiana M. Alves ; Izaura Y. Hirata ; Iuri E. Gouvea ; Marcio F. M. Alves ; Morten Meldal ; Dieter Br&ouml ; mme ; Luiz Juliano ; Maria A. Juliano
• 刊名：Journal of Combinatorial Chemistry
• 出版年：2007
• 出版时间：July 2007
• 年：2007
• 卷：9
• 期：4
• 页码：627 - 634
• 全文大小：448K
• 年卷期：v.9,no.4(July 2007)
• ISSN：1520-4774

文摘

The solubility of peptides in aqueous buffers used for the enzyme assays is a common limitation for allpeptide libraries. In principle, the more water-soluble peptides are, the more susceptible they will be topeptidase hydrolysis. We have demonstrated that this bias can be circumvented in a portion-mixingfluorescence resonance energy transfer (FRET) peptide library by introducing k (lysine in the D-form) inboth termini of the peptides. This more solvated library and another one without the k were assayed usingtrypsin and chymotrypsin as standard peptidases with high selectivity for R and K and for hydrophobic Fand Y, respectively. Significantly improved consistency of the information on substrate profiles was obtainedfrom the solvated library. The influence of improved solvation on substrate specificity determination wassuccessfully demonstrated by the difference in specificity observed between the two libraries employing thehuman cathepsin S (accepts acidic, basic, or neutral amino acids at P₁ position) and Dengue 2 virus NS2B-NS3 protease (high specificity to the pair of basic amino acids K-R, R-R, or Q-R/K at P₂-P₁ positions).In conclusion, hydration of the peptides has a major influence on protease processing, and this bias can bereduced in bound peptide libraries, improving reliability.