Protein–protein cross-reactions involving plastocyanin, cytochrome f and azurin: self-exchange rate constants and related studies with inorganic complexes

Abstract

Protein-protein cross-reactions involving acidic and basic plastocyanins, cytochrome f and azurin, and including reactions between t w o blue copper proteins, have been studied for the first time. The high reactivity of cytochrome f, and properties of the basic Anabaena variabilis plastocyanin, are noted. Reduction potentials for different plastocyanins have been obtained at p H 7-9 and are in the range 340-380 mV. The application of Marcus theory has enabled self-exchange rate constants (25 "C) for acidic parsley (3.3 x l o 3 M-ls-' ) and basic A. variabilis (5.9 x 1 O5 M-l s-l) plastocyanins t o be calculated at pH 7.5, l = 0.1 0 M (NaCI). The influence of overall charge o n the PCu(I)-PCu(II) self-exchange, estimated as -8, -7 in the case of parsley and +1, +2 for A. variabilis, is noted. A mechanism for electron transfer involving contact of adjacent hydrophobic regions on the t w o plastocyanin surfaces giving a Cu 0 Cu separation of 12-14 A is proposed. Rate constants for the cytochrome f(ll) reduction of four PCu(II) plastocyanins (acidic parsley, spinach, Scenedesmus obliquus and basic A. variabilis) are a factor of z 1 O2 less in the case of the basic form. The rate constant for Pseudomonas aeruginosa azurin with cytochrome f has also been determined. The A. variabilis and azurin rate constants give a cytochrome f(ll)-f( Ill) selfexchange of 5.0 x l o 5 M-' s-' at p H 7.5, whereas that obtained from the cross-reaction with the more highly charged parsley PCu(ll) (2.3 x lo8 M-l s-l ) requires correction for work terms, and is predictably out of line. Only one out of four cross-reactions of A. variabilis plastocyanin and azurin with [ C ~( p h e n ) ~] ~+ (phen = 1,lO-phenanthroline) and [Fe(CN)J3-gives a satisfactory fit t o the Marcus theory, indicating an influence of more localised protein charge on these reactions. The absence of a fully developed acidic patch region o n the surface of A. variabilis plastocyanin is discussed. * This value from ref. 35. A value of 5800 M-' cm-' is given in ref. 36.

Spectroscopic and Electrochemical Studies on Active-site Transitions of the Type 1 Copper Protein Pseudoazurin from Achromobacter cycloclastes

“…5). At the lower pH values, the behavior is similar to that observed for plastocyanin (23)(24)(25) and is attributed to the acid-base properties of an active-site histidine of the Cu(I) protein. From the difference between the pK a values for amino acid residues in the two oxidation states of the protein, it is possible to determine, using Equation 1, the effect of protonation or deprotonation of specific amino acid residues on the redox potential of the protein (26):…”

Electron Transfer in Ruthenium-Modified Plastocyanin

“…The unique spectroscopic properties of blue copper include an LMCT [S(Cys) → Cu 2+ ] absorption at ∼600 nm (ε ∼ 4700 M -1 cm -1 ) and a narrow parallel hyperfine EPR splitting pattern. − Plastocyanins have relatively high reduction potentials (340−370 mV vs NHE at pH 7) . The small Cu 2+/+ nuclear reorganization energy and the covalent nature of the Cu−S(Cys) bond facilitate long-range ET reactions with donor and acceptor molecules. ,− …”

Voltammetry of Plastocyanin at a Graphite Electrode: Effects of Structure, Charge, and Electrolyte

“…Figure , reproduced from ref , illustrates the variation in reversible potentials for several plastocyanins under a range of chemical redox conditions. The voltammetric data obtained in the present work, like the data in Figure , also show that positively charged A. variabilis plastocyanin has a considerably more positive E ° f value than any of the negatively charged plastocyanins.…”

“…Antecedent Redox Studies on Plastocyanins. Detailed redox studies on plant plastocyanins by chemical and electrochemical techniques have been reported. − Using rate data derived from chemical redox reactions, Sykes et al. concluded that A. variabilis plastocyanin has a less positive E ° f value than the plastocyanins from two plants (parsley and spinach) and another alga ( Scenedesmus obliquus ).…”

“…Detailed redox studies on plant plastocyanins by chemical and electrochemical techniques have been reported. − Using rate data derived from chemical redox reactions, Sykes et al. concluded that A. variabilis plastocyanin has a less positive E ° f value than the plastocyanins from two plants (parsley and spinach) and another alga ( Scenedesmus obliquus ). Voltammetrically determined E ° f values for four plant plastocyanins (poplar, spinach, cucumber, and parsley) were confined to a narrow range of potentials (389 ± 7 mV vs SHE at 3 °C), although subtle differences among the pH dependences of E ° f and among the electrode−solution interfacial characteristics were found .…”