Document Detail

Pyruvate site of pyruvate phosphate dikinase: crystal structure of the enzyme-phosphonopyruvate complex, and mutant analysis.
MedLine Citation:
PMID:  11790099     Owner:  NLM     Status:  MEDLINE    
Crystals of pyruvate phosphate dikinase in complex with a substrate analogue inhibitor, phosphonopyruvate (K(i) = 3 microM), have been obtained in the presence of Mg(2+). The structure has been determined and refined at 2.2 A resolution, revealing that the Mg(2+)-bound phosphonopyruvate binds in the alpha/beta-barrel's central channel, at the C-termini of the beta-strands. The mode of binding resembles closely the previously proposed PEP substrate binding mode, inferred by the homology of the structure (but not sequence homology) to pyruvate kinase. Kinetic analysis of site-directed mutants, probing residues involved in inhibitor binding, showed that all mutations resulted in inactivation, confirming the key role that these residues play in catalysis. Comparison between the structure of the PPDK-phosphonopyruvate complex and the structures of two complexes of pyruvate kinase, one with Mg(2+)-bound phospholactate and the other with Mg(2+)-oxalate and ATP, revealed that the two enzymes share some key features that facilitate common modes of substrate binding. There are also important structural differences; most notably, the machinery for acid/base catalysis is different.
Osnat Herzberg; Celia C H Chen; Sijiu Liu; Aleksandra Tempczyk; Andrew Howard; Min Wei; Dongmei Ye; Debra Dunaway-Mariano
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Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, Non-P.H.S.; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Biochemistry     Volume:  41     ISSN:  0006-2960     ISO Abbreviation:  Biochemistry     Publication Date:  2002 Jan 
Date Detail:
Created Date:  2002-01-15     Completed Date:  2002-02-13     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  0370623     Medline TA:  Biochemistry     Country:  United States    
Other Details:
Languages:  eng     Pagination:  780-7     Citation Subset:  IM    
Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute, 9600 Gudelsky Drive, Rockville, Maryland 20850, USA.
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MeSH Terms
Amino Acid Substitution
Apoenzymes / chemistry,  metabolism
Binding Sites
Cloning, Molecular
Clostridium / enzymology*
Crystallography, X-Ray
Escherichia coli
Models, Molecular
Mutagenesis, Site-Directed
Protein Conformation
Protein Folding
Protein Structure, Secondary
Pyruvate, Orthophosphate Dikinase / chemistry*,  metabolism
Pyruvic Acid / metabolism*
Recombinant Proteins / chemistry,  metabolism
Grant Support
Reg. No./Substance:
0/Apoenzymes; 0/Recombinant Proteins; 127-17-3/Pyruvic Acid; EC, Orthophosphate Dikinase

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