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Quantification of two-step proteolysis model with consecutive demasking and hydrolysis of peptide bonds using casein hydrolysis by chymotrypsin

Paper ID Volume ID Publish Year Pages File Format Full-Text
3402 168 2013 9 PDF Available
Title
Quantification of two-step proteolysis model with consecutive demasking and hydrolysis of peptide bonds using casein hydrolysis by chymotrypsin
Abstract

•We model demasking and hydrolysis of peptide bonds during proteolysis.•Demasked bonds are hydrolysed with different hydrolysis rate constants.•The hydrolysis of casein by chymotrypsin is analyzed.•Demasking rate constant is lower than hydrolysis rate constant for PheX bonds.•A half of peptide bonds is initially masked in casein.

Enzymatic hydrolysis of peptide bonds becomes possible after removing steric obstacles shielding polypeptide sites against enzymatic attack, i.e. after demasking of these sites. In a simple two-step model, proteolysis was regarded as a two-step process with consecutive demasking and hydrolysis stages. A new analytical procedure was suggested to determine three experimental kinetic parameters: demasking rate constant kd, degree of initially masked peptide bonds m and maximum hydrolysis rate constant kh. The approach was shown on the example of the hydrolysis of summary casein by chymotrypsin (25 °C, pH 7.5). Kinetic analysis includes the determination of an apparent Michaelis constant that we regard as a function of the degree of peptide bond hydrolysis. Two sets of hydrolysis rate constants were used to calculate the parameters of a two-step model. It was found that kd is lower than the hydrolysis rate constants for specific sites consisting of aromatic amino acid residues, and at least a half of peptide bonds is initially masked in casein. Using parameters of a two-step model, we calculated second-order rate constant and degrees of hydrolysis for specific peptide bonds as functions of the hydrolysis degree.

Keywords
Proteolysis model; Chymotryptic digestion; Enzymatic kinetics; Peptide bond demasking; Casein hydrolyzates
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Quantification of two-step proteolysis model with consecutive demasking and hydrolysis of peptide bonds using casein hydrolysis by chymotrypsin
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Publisher
Database: Elsevier - ScienceDirect
Journal: Biochemical Engineering Journal - Volume 74, 15 May 2013, Pages 60–68
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us