Abstract |
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 P2-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.
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Authors | Fabiana M Alves, Izaura Y Hirata, Iuri E Gouvea, Marcio F M Alves, Morten Meldal, Dieter Brömme, Luiz Juliano, Maria A Juliano |
Journal | Journal of combinatorial chemistry
(J Comb Chem)
2007 Jul-Aug
Vol. 9
Issue 4
Pg. 627-34
ISSN: 1520-4766 [Print] United States |
PMID | 17563123
(Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)
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Chemical References |
- Cross-Linking Reagents
- NS2B protein, flavivirus
- Peptide Library
- Peptides
- Solutions
- Viral Nonstructural Proteins
- Polyethylene Glycols
- Cathepsins
- NS3 protease, dengue virus
- Serine Endopeptidases
- cathepsin S
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Topics |
- Amino Acid Sequence
- Cathepsins
(metabolism)
- Cross-Linking Reagents
(chemistry)
- Dengue Virus
(enzymology)
- Fluorescence Resonance Energy Transfer
- Humans
- Kinetics
- Peptide Library
- Peptides
(chemistry, metabolism)
- Polyethylene Glycols
(chemistry)
- Serine Endopeptidases
(metabolism)
- Solutions
(chemistry)
- Substrate Specificity
- Viral Nonstructural Proteins
(metabolism)
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