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Journal article
Design of a Ruthenium-Cytochrome c Derivative to Measure Electron Transfer to the Initial Acceptor in Cytochrome c Oxidase
The Journal of biological chemistry, v 270(6), pp 2466-2472
10 Feb 1995
PMID: 7852307
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Abstract
A ruthenium-labeled cytochrome c derivative was prepared to meet two design criteria: the ruthenium group must transfer an electron rapidly to the heme group, but not alter the interaction with cytochrome c oxidase. Site-directed mutagenesis was used to replace His39 on the backside of yeast C102T iso-1-cytochrome c with a cysteine residue, and the single sulfhydryl group was labeled with (4-bromomethyl-4′methylbipyridine) (bisbipyridine)ruthenium(II) to form Ru-39-cytochrome c (cyt c). There is an efficient pathway for electron transfer from the ruthenium group to the heme group of Ru-39-cyt c comprising 13 covalent bonds and one hydrogen bond. Electron transfer from the excited state Ru(II∗) to ferric heme c occurred with a rate constant of (6.0 ± 2.0) × 105 s−1, followed by electron transfer from ferrous heme c to Ru(III) with a rate constant of (1.0 ± 0.2) × 106 s−1. Laser excitation of a complex between Ru-39-cyt c and beef cytochrome c oxidase in low ionic strength buffer (5 mM phosphate, pH 7) resulted in electron transfer from photoreduced heme c to CuA with a rate constant of (6 ± 2) × 104 s−1, followed by electron transfer from CuA to heme a with a rate constant of (1.8 ± 0.3) × 104 s−1. Increasing the ionic strength to 100 mM leads to bimolecular kinetics as the complex is dissociated. The second-order rate constant is (2.5 ± 0.4) × 107M−1 s−1 at 230 mM ionic strength, nearly the same as that of wild-type iso-1-cytochrome c.
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Details
- Title
- Design of a Ruthenium-Cytochrome c Derivative to Measure Electron Transfer to the Initial Acceptor in Cytochrome c Oxidase
- Creators
- Lois M. Geren - University of Arkansas at FayettevilleJames R. Beasley - University of North Carolina at Chapel HillBryan R. Fine - University of North Carolina at Chapel HillAleister J. Saunders - University of North Carolina at Chapel HillSharon Hibdon - University of Arkansas at FayettevilleGary J. Pielak - University of North Carolina at Chapel HillBill Durham - University of Arkansas at FayettevilleFrancis Millett - University of Arkansas at Fayetteville
- Publication Details
- The Journal of biological chemistry, v 270(6), pp 2466-2472
- Publisher
- Elsevier
- Number of pages
- 7
- Grant note
- This work was supported in part by National Institutes of Health Grants GM20488 (to F. M. and B. D.) and GM42501 (to G. J. P.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- Biology
- Web of Science ID
- WOS:A1995QF53500013
- Scopus ID
- 2-s2.0-0028819233
- Other Identifier
- 991021448037504721
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- Collaboration types
- Domestic collaboration
- Web of Science research areas
- Biochemistry & Molecular Biology