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rdf:type |
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lifeskim:mentions |
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pubmed:issue |
2
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pubmed:dateCreated |
1996-9-10
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pubmed:abstractText |
This paper reports the biochemical characterization of the flavoprotein L-aspartate oxidase from Escherichia coli. Modification of a previously published procedure allowed overexpression of the holoenzyme in an unproteolysed form. L-Aspartate oxidase is a monomer of 60 kDa containing 1 mol of noncovalently bound FAD/mol protein. A polarographic and two spectrophotometric coupled assays have been set up to monitor the enzymatic activity continuously. L-Aspartate oxidase was subjected to product inhibition since iminoaspartate, which results from the oxidation of L-aspartate, binds to the enzyme with a dissociation constant (Kd) equal to 1.4 microM. The enzyme binds FAD by a simple second-order process with Kd 0.67 microM. Site-directed mutagenesis of the residues E43, G44, S45, F47 and Y48 located in the putative binding site of the isoallossazinic portion of FAD reduces the affinity for the coenzyme.
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pubmed:language |
eng
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pubmed:journal |
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pubmed:citationSubset |
IM
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pubmed:chemical |
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pubmed:status |
MEDLINE
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pubmed:month |
Jul
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pubmed:issn |
0014-2956
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pubmed:author |
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pubmed:issnType |
Print
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pubmed:day |
15
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pubmed:volume |
239
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
418-26
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pubmed:dateRevised |
2007-7-23
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pubmed:meshHeading |
pubmed-meshheading:8706749-Amino Acid Oxidoreductases,
pubmed-meshheading:8706749-Amino Acid Sequence,
pubmed-meshheading:8706749-Bacillus subtilis,
pubmed-meshheading:8706749-Base Sequence,
pubmed-meshheading:8706749-Binding Sites,
pubmed-meshheading:8706749-Conserved Sequence,
pubmed-meshheading:8706749-Escherichia coli,
pubmed-meshheading:8706749-Flavin-Adenine Dinucleotide,
pubmed-meshheading:8706749-Genes, Bacterial,
pubmed-meshheading:8706749-Isoenzymes,
pubmed-meshheading:8706749-Kinetics,
pubmed-meshheading:8706749-Molecular Sequence Data,
pubmed-meshheading:8706749-Mutagenesis, Site-Directed,
pubmed-meshheading:8706749-Oligodeoxyribonucleotides,
pubmed-meshheading:8706749-Point Mutation,
pubmed-meshheading:8706749-Recombinant Proteins,
pubmed-meshheading:8706749-Sequence Homology, Amino Acid,
pubmed-meshheading:8706749-Spectrophotometry
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pubmed:year |
1996
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pubmed:articleTitle |
L-aspartate oxidase from Escherichia coli. I. Characterization of coenzyme binding and product inhibition.
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pubmed:affiliation |
Istituto di Fisiologia Veterinaria e Biochimica, Università di Milano, Italy.
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pubmed:publicationType |
Journal Article,
Comparative Study,
Research Support, Non-U.S. Gov't
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