TY - JOUR

T1 - Controlled peptide solvation in portion-mixing libraries of FRET peptides

T2 - Improved specificity determination for dengue 2 virus NS2B-NS3 protease and human cathepsin S

AU - Alves, Fabiana M.

AU - Hirata, Izaura Y.

AU - Gouvea, Iuri E.

AU - Alves, Mareio F.M.

AU - Meldal, Morten

AU - Brömme, Dieter

AU - Juliano, Luiz

AU - Juliano, Maria A.

PY - 2007/7

Y1 - 2007/7

N2 - The solubility of peptides in aqueous buffers used for the enzyme assays is a common limitation for all peptide libraries. In principle, the more water-soluble peptides are, the more susceptible they will be to peptidase hydrolysis. We have demonstrated that this bias can be circumvented in a portion-mixing fluorescence resonance energy transfer (FRET) peptide library by introducing k (lysine in the D-form) in both termini of the peptides. This more solvated library and another one without the k were assayed using trypsin and chymotrypsin as standard peptidases with high selectivity for R and K and for hydrophobic F and Y, respectively. Significantly improved consistency of the information on substrate profiles was obtained from the solvated library. The influence of improved solvation on substrate specificity determination was successfully demonstrated by the difference in specificity observed between the two libraries employing the human 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 P 2-P1 positions). In conclusion, hydration of the peptides has a major influence on protease processing, and this bias can be reduced in bound peptide libraries, improving reliability.

AB - The solubility of peptides in aqueous buffers used for the enzyme assays is a common limitation for all peptide libraries. In principle, the more water-soluble peptides are, the more susceptible they will be to peptidase hydrolysis. We have demonstrated that this bias can be circumvented in a portion-mixing fluorescence resonance energy transfer (FRET) peptide library by introducing k (lysine in the D-form) in both termini of the peptides. This more solvated library and another one without the k were assayed using trypsin and chymotrypsin as standard peptidases with high selectivity for R and K and for hydrophobic F and Y, respectively. Significantly improved consistency of the information on substrate profiles was obtained from the solvated library. The influence of improved solvation on substrate specificity determination was successfully demonstrated by the difference in specificity observed between the two libraries employing the human 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 P 2-P1 positions). In conclusion, hydration of the peptides has a major influence on protease processing, and this bias can be reduced in bound peptide libraries, improving reliability.

UR - http://www.scopus.com/inward/record.url?scp=34547218898&partnerID=8YFLogxK

U2 - 10.1021/cc070042k

DO - 10.1021/cc070042k

M3 - Journal article

C2 - 17563123

AN - SCOPUS:34547218898

VL - 9

SP - 627

EP - 634

JO - ACS Combinatorial Science

JF - ACS Combinatorial Science

SN - 2156-8952

IS - 4

ER -