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Understanding the structure–function role of specific catalytic residues in a model food related enzyme: Pepsin

Paper ID Volume ID Publish Year Pages File Format Full-Text
18160 42713 2007 6 PDF Available
Title
Understanding the structure–function role of specific catalytic residues in a model food related enzyme: Pepsin
Abstract

Several aspartic proteinases are used as food processing aids (e.g., pepsin and chymosin in the manufacture of cheese). Aspartic proteinases contain two active site catalytic aspartic acid residues that act as an acid–base pair. Pepsin active site glutamic acid mutants D32E, D215E, and D32E/D215E were compared to wild-type. All mutants were unable to activate autocatalytically, but were able to retain activity and showed decreased turnover numbers (kcat), as well as decreased catalytic efficiencies (kcat/Km). The D32E substitution resulted in a decrease in activity at pH 1–2 suggesting a change in the pKa of the normally conserved residue 32. Circular dichroism revealed that overall secondary structure contents were unaffected by catalytic glutamic acid substitutions whereas thermal stabilities (Tm) were lower relative to wild-type. Energy minimizations predicted that glutamic acid at position 32 altered the positioning of the catalytic carboxyl group more than position 215. This study shows that pepsin can function as a glutamic proteinase.

Keywords
Active site; Denaturation; Model; Mutation; Pepsin
First Page Preview
Understanding the structure–function role of specific catalytic residues in a model food related enzyme: Pepsin
Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 40, Issue 5, 3 April 2007, Pages 1175–1180
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering