Document Detail


Site-directed mutagenesis reveals functional contribution of Thr218, Lys220 and Asp304 in chymosin.
MedLine Citation:
PMID:  2290836     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The functional contributions of amino acid residues Thr218 and Asp304 of chymosin, both of which are highly conserved in the aspartic proteinases, are analysed by means of site-directed mutagenesis. The optimum pH values, milk-clotting (C) and proteolytic (P) activities and kinetic parameters for synthetic oligopeptides as substrates were examined for the mutant enzymes. The mutation Thr218Ser caused a marked increase in the C/P ratio, which seemed to be due to a change in substrate recognition. Although the negative charge of Asp304 had been expected to play a role in lowering the optimum pH values in the aspartic proteinases, this turned out not to be the case in chymosin because both the mutations Asp304Ala and Asp304Glu caused a similar shift of the optimum pH towards the acidic side. In addition, the mutation Lys220Leu, which we generated previously, was found to cause a decrease in the C/P ratio, mainly due to the increase in the proteolytic activity.
Authors:
J Suzuki; A Hamu; M Nishiyama; S Horinouchi; T Beppu
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Protein engineering     Volume:  4     ISSN:  0269-2139     ISO Abbreviation:  Protein Eng.     Publication Date:  1990 Oct 
Date Detail:
Created Date:  1991-04-03     Completed Date:  1991-04-03     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  8801484     Medline TA:  Protein Eng     Country:  ENGLAND    
Other Details:
Languages:  eng     Pagination:  69-71     Citation Subset:  IM    
Affiliation:
Department of Agricultural Chemistry, Faculty of Agriculture, University of Tokyo, Japan.
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MeSH Terms
Descriptor/Qualifier:
Amino Acid Sequence
Base Sequence
Chymosin / chemistry,  genetics*,  metabolism
Escherichia coli / enzymology*
Hydrogen-Ion Concentration
Kinetics
Molecular Sequence Data
Mutagenesis, Site-Directed
Protein Engineering
Substrate Specificity
Chemical
Reg. No./Substance:
EC 3.4.23.4/Chymosin

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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