The path of proteolysis by bovine chymotrypsin.

Autor: Vreeke GJC; Laboratory of Food Chemistry, Wageningen University & Research, P.O. 17, 6708 AA Wageningen, the Netherlands., Vincken JP; Laboratory of Food Chemistry, Wageningen University & Research, P.O. 17, 6708 AA Wageningen, the Netherlands., Wierenga PA; Laboratory of Food Chemistry, Wageningen University & Research, P.O. 17, 6708 AA Wageningen, the Netherlands. Electronic address: peter.wierenga@wur.nl.
Jazyk: angličtina
Zdroj: Food research international (Ottawa, Ont.) [Food Res Int] 2023 Mar; Vol. 165, pp. 112485. Date of Electronic Publication: 2023 Jan 21.
DOI: 10.1016/j.foodres.2023.112485
Abstrakt: Chymotrypsin is one of the major proteases in intestinal protein digestion. Observations about the type of bonds that are hydrolysed (specificity and preference) were in the past derived from the peptide composition after digestion or hydrolysis rates of synthetic peptides. In this study, the path of hydrolysis by bovine chymotrypsin, i.e formation and degradation of peptides, were described for α-lactalbumin, β-lactoglobulin and β-casein. The peptide compositions, determined with UPLC-PDA-MS at different time points were used to determine the digestion kinetics for individual cleavage sites. It was evaluated how statements on (secondary) specificity from literature were reflected in the release kinetics of peptides. β-Lactoglobulin reached the highest degree of hydrolysis (10.9 ± 0.1 %) and was hydrolysed fastest (28 ± 1 mM peptide bonds /s/mM enzyme ), regardless of its globular (tertiary) structure. Chymotrypsin showed a preference towards aromatic amino acids, methionine and leucine, but was also tolerant to other amino acids. For the cleavage sites within this preference, ̴73% of the cleavage sites were hydrolysed with high or intermediate selectivity. For the missed cleavages within the preference, 45 % was explained by hindrance of proline, which affected hydrolysis only when in positions P3, P1' or P2'. No clear indication (based on primary structure) was found to explain the other missed cleavages. A few cleavage sites were hydrolysed extremely efficient in α-lactalbumin (F9, F31, W104) and β-casein (W143, L163, F190). This study gave unique and quantitative insight in peptide formation and degradation by chymotrypsin in the digestion of proteins. The approach used showed potential to explore the path of hydrolysis for other proteases with less defined specificity.
Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)
Databáze: MEDLINE
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