Multichannel Analysis of Single-Turnover Kinetics of Cytochrome aa3 Reduction of O2
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- 作者：Salil Bose ; Richard W. Hendler ; Richard I. Shrager ; Sunney I. Chan ; and Paul D. Smith
- 刊名：Biochemistry
- 出版年：1997
- 出版时间：March 4, 1997
- 年：1997
- 卷：36
- 期：9
- 页码：2439 - 2449
- 全文大小：227K
- 年卷期：v.36,no.9(March 4, 1997)
- ISSN：1520-4995
文摘
The single-turnover kinetics of the oxidation of cytochromeaa3 by O2 have beenstudiedusing a new approach. Up to 1000 whole spectra covering both theSoret and 
regions were sequentiallycollected at room temperature from single samples with a timeresolution of 10 
s. All of the spectraland time information were used in analyses based on singular valuedecomposition. Four spectral transitions(i.e., intermediates) were distinguished with time constants near 0.01,0.1, 1.1, and 30 ms. Two differentkinds of sequential models were evaluated, one linear and the otherbranched. Although past kineticanalyses have emphasized the linear sequential model, the complexity ofthe intramolecular electron transferin this enzyme suggests that a branched model be considered. Thisis especially true in a single-turnoverexperiment where earlier optical and EPR studies have pointedunequivocally to a branched model [Cloreet al. (1980) Biochem. J. 185, 139-154; Blairet al. (1985) J. Am. Chem. Soc. 107,7389-7399]. In thepresent study, analysis of spectral data in terms of the linear modeldid not reveal the formation anddecay of the expected oxyferryl intermediate, whereas analysis of thebranched model did. The resultsobtained using the branched model are consistent with all of theavailable evidence from a broad rangeof physical techniques that have been applied to examine thesingle-turnover kinetics of the oxidation ofreduced cytochrome aa3 byO2.
regions were sequentiallycollected at room temperature from single samples with a timeresolution of 10 s. All of the spectraland time information were used in analyses based on singular valuedecomposition. Four spectral transitions(i.e., intermediates) were distinguished with time constants near 0.01,0.1, 1.1, and 30 ms. Two differentkinds of sequential models were evaluated, one linear and the otherbranched. Although past kineticanalyses have emphasized the linear sequential model, the complexity ofthe intramolecular electron transferin this enzyme suggests that a branched model be considered. Thisis especially true in a single-turnoverexperiment where earlier optical and EPR studies have pointedunequivocally to a branched model [Cloreet al. (1980) Biochem. J. 185, 139-154; Blairet al. (1985) J. Am. Chem. Soc. 107,7389-7399]. In thepresent study, analysis of spectral data in terms of the linear modeldid not reveal the formation anddecay of the expected oxyferryl intermediate, whereas analysis of thebranched model did. The resultsobtained using the branched model are consistent with all of theavailable evidence from a broad rangeof physical techniques that have been applied to examine thesingle-turnover kinetics of the oxidation ofreduced cytochrome aa3 byO2.