Source:http://linkedlifedata.com/resource/pubmed/id/11591146
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rdf:type | |
lifeskim:mentions | |
pubmed:issue |
41
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pubmed:dateCreated |
2001-10-9
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pubmed:abstractText |
Glutamate 47 is conserved in 1-aminocyclopropane-1-carboxylate (ACC) synthases and is positioned near the sulfonium pole of (S,S)-S-adenosyl-L-methionine (SAM) in the modeled pyridoxal phosphate quinonoid complex with SAM. E47Q and E47D constructs of ACC synthase were made to investigate a putative ionic interaction between Glu47 and SAM. The k(cat)/K(m) values for the conversion of (S,S)-SAM to ACC and methylthioadenosine (MTA) are depressed 630- and 25-fold for the E47Q and E47D enzymes, respectively. The decreases in the specificity constants are due to reductions in k(cat) for both mutant enzymes, and a 5-fold increase in K(m) for the E47Q enzyme. Importantly, much smaller effects were observed for the kinetic parameters of reactions with the alternate substrates L-vinylglycine (L-VG) (deamination to form alpha-ketobutyrate and ammonia) and L-alanine (transamination to form pyruvate), which have uncharged side chains. L-VG is both a substrate and a mechanism-based inactivator of the enzyme [Feng, L., and Kirsch, J. F. (2000) Biochemistry 39, 2436-2444], but the partition ratio, k(cat)/k(inact), is unaffected by the Glu47 mutations. ACC synthase primarily catalyzes the beta,gamma-elimination of MTA from the (R,S) diastereomer of SAM to produce L-VG [Satoh, S., and Yang, S. F. (1989) Arch.Biochem. Biophys. 271, 107-112], but catalyzes the formation of ACC to a lesser extent via alpha,gamma-elimination of MTA. The partition ratios for (alpha,gamma/beta,gamma)-elimination on (R,S)-SAM are 0.4, < or =0.014, and < or =0.08 for the wild-type, E47Q, and E47D enzymes, respectively. The results of these experiments strongly support a role for Glu47 as an anchor for the sulfonium pole of (S,S)-SAM, and consequently a role as an active site determinant of reaction specificity.
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pubmed:grant | |
pubmed:language |
eng
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pubmed:journal | |
pubmed:citationSubset |
IM
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pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/1-aminocyclopropanecarboxylate...,
http://linkedlifedata.com/resource/pubmed/chemical/Glutamic Acid,
http://linkedlifedata.com/resource/pubmed/chemical/Lyases,
http://linkedlifedata.com/resource/pubmed/chemical/Recombinant Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/S-Adenosylmethionine
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pubmed:status |
MEDLINE
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pubmed:month |
Oct
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pubmed:issn |
0006-2960
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pubmed:author | |
pubmed:issnType |
Print
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pubmed:day |
16
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pubmed:volume |
40
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
12276-84
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pubmed:dateRevised |
2007-11-14
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pubmed:meshHeading |
pubmed-meshheading:11591146-Catalytic Domain,
pubmed-meshheading:11591146-Glutamic Acid,
pubmed-meshheading:11591146-Kinetics,
pubmed-meshheading:11591146-Lyases,
pubmed-meshheading:11591146-Models, Molecular,
pubmed-meshheading:11591146-Mutagenesis, Site-Directed,
pubmed-meshheading:11591146-Plants,
pubmed-meshheading:11591146-Recombinant Proteins,
pubmed-meshheading:11591146-S-Adenosylmethionine,
pubmed-meshheading:11591146-Substrate Specificity,
pubmed-meshheading:11591146-Viscosity
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pubmed:year |
2001
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pubmed:articleTitle |
Glutamate 47 in 1-aminocyclopropane-1-carboxylate synthase is a major specificity determinant.
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pubmed:affiliation |
Department of Molecular and Cell Biology, Division of Biochemistry and Molecular Biology, 229 Stanley Hall, University of California, Berkeley, 94720-3206, USA.
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pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.,
Research Support, U.S. Gov't, Non-P.H.S.,
Research Support, Non-U.S. Gov't
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