Source:http://linkedlifedata.com/resource/pubmed/id/12069606
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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
25
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| pubmed:dateCreated |
2002-6-18
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| pubmed:abstractText |
Residues glutamate 156 (E156), aspartate 190 (D190), asparagine 181 (N181), lysine 186 (K186), and asparagine 191 (N191) in the active site of S-adenosylhomocysteine (AdoHcy) hydrolase have been mutated to alanine (A). AdoHcy hydrolase achieves catalysis of AdoHcy hydrolysis to adenosine (Ado) and homocysteine (Hcy) by means of a redox partial reaction (3'-oxidation of AdoHcy at the beginning and 3'-reduction of Ado at the end of the catalytic cycle) spanning an elimination/addition partial reaction (elimination of Hcy from the oxidized substrate and addition of water to generate the oxidized product), with the enzyme in an open NAD(+) form in the ligand-free state and in a closed NADH form during the elimination/addition partial reaction. Mutation K186A reduces the rate of a model enzymatic reaction for the redox partial reaction by a factor of 280000 and the rate of a model reaction for the elimination/addition partial reaction by a factor of 24000, consistent with a primary catalytic role in both partial reactions as a proton donor/acceptor at the 3'-OH/3'-keto center. Secondary roles for N181 and N191 in localizing the flexible side chain of K186 in a catalytically effective position are supported by rate reduction factors for N181A of 2500 (redox) and 240 (elimination/addition) and for N191A of 730 (redox) and 340 (elimination/addition). A role of D190 in orienting the substrate for effective transition-state stabilization is consistent with rate reduction factors of 1300 (redox) and 30 (elimination/addition) for D190A. Residue E156 may act to maintain K186 in the desired protonation state: rate deduction factors are 1100 (redox) and 70 (elimination/addition). The mutational increases in free energy barriers for k(cat)/K(M) are described by a linear combination of the effects for the partial reactions with the coefficients equal to the fractional degree that each partial reaction determines the rate for k(cat)/K(M). A similar linear equation for k(cat) overestimates the barrier increase by a uniform 5 kJ/mol, probably reflecting reactant-state stabilization by the wild-type enzyme that is abolished by the mutations.
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| pubmed:grant | |
| pubmed:language |
eng
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| pubmed:journal | |
| pubmed:citationSubset |
IM
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| pubmed:chemical | |
| pubmed:status |
MEDLINE
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| pubmed:month |
Jun
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| pubmed:issn |
0006-2960
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| pubmed:author | |
| pubmed:issnType |
Print
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| pubmed:day |
25
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| pubmed:volume |
41
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| pubmed:owner |
NLM
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| pubmed:authorsComplete |
Y
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| pubmed:pagination |
8134-42
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| pubmed:dateRevised |
2007-11-14
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| pubmed:meshHeading |
pubmed-meshheading:12069606-Adenosylhomocysteinase,
pubmed-meshheading:12069606-Binding Sites,
pubmed-meshheading:12069606-Catalysis,
pubmed-meshheading:12069606-Humans,
pubmed-meshheading:12069606-Hydrolases,
pubmed-meshheading:12069606-Models, Chemical,
pubmed-meshheading:12069606-Mutagenesis, Site-Directed,
pubmed-meshheading:12069606-Oxidation-Reduction,
pubmed-meshheading:12069606-Point Mutation,
pubmed-meshheading:12069606-Protein Structure, Quaternary,
pubmed-meshheading:12069606-Protein Structure, Secondary,
pubmed-meshheading:12069606-Recombinant Proteins,
pubmed-meshheading:12069606-Thermodynamics
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| pubmed:year |
2002
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| pubmed:articleTitle |
Contributions of active site residues to the partial and overall catalytic activities of human S-adenosylhomocysteine hydrolase.
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| pubmed:affiliation |
Departments of Molecular Biosciences and Pharmaceutical Chemistry, Simons Research Laboratories, The University of Kansas, Lawrence, KS 66047, USA.
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| pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.
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