title: Characterisation of the electron self-exchange rates in hexametaphosphate-cytochrome-c aggregates measured using high-resolution 1H-NMR spectroscopy.
abstract: 1H-NMR spectroscopy has been used to measure the rate of unimolecular electron exchange between cytochrome c molecules in protein aggregates stabilised by the addition of sodium hexametaphosphate. The average intracomplex electron exchange rate is measured from line broadening of hyperfine-shifted resonances of ferricytochrome c in an equimolar mixture of reduced and oxidised protein. The line-broadening due to electron exchange is significantly greater than that due to protein aggregation and reaches a maximum value between 1-2 mol hexametaphosphate/mol protein. Significantly the exchange-induced broadening is a first-order process and is directly proportional to the size of the cytochrome c oligomer. From the temperature dependence of exchange broadening the activation enthalpy was estimated to be 75.8 kJ mol-1 whereas the activation entropy was 295 J mol-1 K-1 for a dimer of cytochrome c at a hexametaphosphate/protein molar ratio of 1. Both activation parameters decrease in magnitude as the order of the cytochrome c oligomer increases. The rates of intracomplex electron exchange in Saccharomyces cerevisiae iso-2 and Candida krusei cytochromes c are lower than that of the horse protein, implying that primary sequence plays a fundamental part in determining the rate of exchange. The relevance of these observations is discussed in terms of the function of cytochrome c.
authors.affiliation: Inorganic Chemistry Laboratory, University of Oxford, England.
authors.forename: D W
authors.initials: DW
authors.lastname: Concar
authors.forename: D
authors.initials: D
authors.lastname: Whitford
authors.forename: R J
authors.initials: RJ
authors.lastname: Williams
mesh_terms: D000818:Animals; D001665:Binding Sites; D002151:Calorimetry; D002175:Candida / Q000378:metabolism; D003574:Cytochrome c Group / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006736:Horses; D006859:Hydrogen; D007700:Kinetics; D046911:Macromolecular Substances; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D009206:Myocardium / Q000378:metabolism; D010710:Phosphates / Q000378:metabolism*
chemical_list: D003574:Cytochrome c Group; D046911:Macromolecular Substances; D010710:Phosphates; C005035:hexametaphosphate; D006859:Hydrogen