Source:http://linkedlifedata.com/resource/pubmed/id/10074352
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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
10
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| pubmed:dateCreated |
1999-3-23
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| pubmed:abstractText |
Recent studies [Mallett, T. C., and Claiborne, A. (1998) Biochemistry 37, 8790-8802] of the O2 reactivity of C42S NADH oxidase (O2 --> H2O2) revealed an asymmetric mechanism in which the two FADH2.NAD+ per reduced dimer display kinetic inequivalence. In this report we provide evidence indicating that the fully active, recombinant wild-type oxidase (O2 --> 2H2O) displays thermodynamic inequivalence between the two active sites per dimer. Using NADPH to generate the free reduced wild-type enzyme (EH2'/EH4), we have shown that NAD+ titrations lead to differential behavior as only one FADH2 per dimer binds NAD+ tightly to give the charge-transfer complex. The second FADH2, in contrast, transfers its electrons to the single Cys42-sulfenic acid (Cys42-SOH) redox center, which remains oxidized during the reductive titration. Titrations of the reduced NADH oxidase with oxidized 3-acetylpyridine and 3-aminopyridine adenine dinucleotides further support the conclusion that the two FADH2 per dimer in wild-type enzyme can be described as distinct "charge-transfer" and "electron-transfer" sites, with the latter site giving rise to either intramolecular (Cys42-SOH) or bimolecular (pyridine nucleotide) reduction. The reduced C42S mutant is not capable of intramolecular electron transfer on binding pyridine nucleotides, thus confirming that the Cys42-SOH center is in fact the source of the redox asymmetry observed with wild-type oxidase. These observations on the role of Cys42-SOH in the expression of thermodynamic inequivalence as observed in wild-type NADH oxidase complement the previously described kinetic inequivalence of the C42S mutant; taken together, these results provide the overlapping framework for an alternating sites cooperativity model of oxidase action.
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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/3-acetylpyridine adenine...,
http://linkedlifedata.com/resource/pubmed/chemical/Bacterial Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/Coenzymes,
http://linkedlifedata.com/resource/pubmed/chemical/Cysteine,
http://linkedlifedata.com/resource/pubmed/chemical/Dithionite,
http://linkedlifedata.com/resource/pubmed/chemical/Multienzyme Complexes,
http://linkedlifedata.com/resource/pubmed/chemical/NAD,
http://linkedlifedata.com/resource/pubmed/chemical/NADH, NADPH Oxidoreductases,
http://linkedlifedata.com/resource/pubmed/chemical/NADH oxidase,
http://linkedlifedata.com/resource/pubmed/chemical/NADP,
http://linkedlifedata.com/resource/pubmed/chemical/Recombinant Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/Serine,
http://linkedlifedata.com/resource/pubmed/chemical/Sulfenic Acids
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| pubmed:status |
MEDLINE
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| pubmed:month |
Mar
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| pubmed:issn |
0006-2960
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| pubmed:author | |
| pubmed:issnType |
Print
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| pubmed:day |
9
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| pubmed:volume |
38
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| pubmed:owner |
NLM
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| pubmed:authorsComplete |
Y
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| pubmed:pagination |
3000-11
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| pubmed:dateRevised |
2007-11-14
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| pubmed:meshHeading |
pubmed-meshheading:10074352-Bacterial Proteins,
pubmed-meshheading:10074352-Binding Sites,
pubmed-meshheading:10074352-Coenzymes,
pubmed-meshheading:10074352-Cysteine,
pubmed-meshheading:10074352-Dithionite,
pubmed-meshheading:10074352-Electron Transport,
pubmed-meshheading:10074352-Enzyme Stability,
pubmed-meshheading:10074352-Hydrogen-Ion Concentration,
pubmed-meshheading:10074352-Multienzyme Complexes,
pubmed-meshheading:10074352-Mutagenesis, Site-Directed,
pubmed-meshheading:10074352-NAD,
pubmed-meshheading:10074352-NADH, NADPH Oxidoreductases,
pubmed-meshheading:10074352-NADP,
pubmed-meshheading:10074352-Oxidation-Reduction,
pubmed-meshheading:10074352-Recombinant Proteins,
pubmed-meshheading:10074352-Serine,
pubmed-meshheading:10074352-Spectrometry, Fluorescence,
pubmed-meshheading:10074352-Sulfenic Acids
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| pubmed:year |
1999
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| pubmed:articleTitle |
Equilibrium analyses of the active-site asymmetry in enterococcal NADH oxidase: role of the cysteine-sulfenic acid redox center.
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| pubmed:affiliation |
Department of Biochemistry, Wake Forest University Medical Center, Winston-Salem, North Carolina 27157, USA.
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| pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.
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