Document Detail


The glycine-rich region of Escherichia coli D-serine dehydratase. Altered interactions with pyridoxal 5'-phosphate produced by substitution of aspartic acid for glycine.
MedLine Citation:
PMID:  3053700     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Replacement of glycine by aspartic acid at either of two sites in a conserved, glycine-rich region inactivates the pyridoxal 5'-phosphate-dependent enzyme D-serine dehydratase (DSD) from Escherichia coli. To investigate why aspartic acid at position 279 or 281 causes a loss of activity, we measured the affinity of the G----D variants for pyridoxal 5'-phosphate and a cofactor:substrate analog complex and compared the UV, CD, and fluorescence properties of wild-type D-serine dehydratase and the inactive variants. The two G----D variants DSD(G279D) and DSD (G281D) displayed marked differences from wild-type D-serine dehydratase and from each other with respect to their affinity for pyridoxal 5'-phosphate and for a pyridoxal 5'-phosphate:glycine Schiff base. Compared to the wild-type enzyme, the cofactor affinity of DSD(G279D) and DSD(G281D) was decreased 225- and 50-fold, respectively, and the ability to retain a cofactor:glycine complex was decreased 765- and 1970-fold. The spectral properties of the inactive variants suggest that they form a Schiff base linkage with pyridoxal 5'-phosphate but do not hold the cofactor in a catalytically competent orientation. Moreover, the amount of cofactor aldamine in equilibrium with cofactor Schiff base is increased in DSD(G279D) and DSD(G281D) relative to that in wild-type DSD. Collectively, our findings indicate that introduction of a carboxymethyl side chain at G-279 or G-281 directly or indirectly disrupts catalytically essential protein-cofactor and protein-substrate interactions and thereby prevents processing of the enzyme bound cofactor:substrate complex. The conserved glycine-rich region is thus either an integral part of the D-serine dehydratase active site or conformationally linked to it.
Authors:
M Marceau; S D Lewis; J A Shafer
Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  263     ISSN:  0021-9258     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  1988 Nov 
Date Detail:
Created Date:  1988-12-14     Completed Date:  1988-12-14     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  16934-41     Citation Subset:  IM    
Affiliation:
Department of Biological Chemistry, University of Michigan, Ann Arbor 48109.
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MeSH Terms
Descriptor/Qualifier:
Algorithms
Aspartic Acid / metabolism*
Circular Dichroism
Escherichia coli / enzymology*
Glycine / analysis*
L-Serine Dehydratase / analysis,  metabolism*
Pyridoxal Phosphate / metabolism*
Structure-Activity Relationship
Grant Support
ID/Acronym/Agency:
HL32006/HL/NHLBI NIH HHS
Chemical
Reg. No./Substance:
54-47-7/Pyridoxal Phosphate; 56-40-6/Glycine; 56-84-8/Aspartic Acid; EC 4.3.1.17/L-Serine Dehydratase

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


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