Ruthenated proteins, electron transfer

Rate constants, k, are for electron transfer from the donor to acceptor groups in ruthenated electron transfer proteins. Ruthenium ligands are abbreviated as follows NH3, a isonicotinamide, isn pyridine, py. MP are metallosubstituted porphyrins (M = Fe, Zn, Pd, Pt, Mg, Cd, and H), MP and MP- + are the photoexcited and oxidized radical cation forms of MP, respectively. PDE is porphyrin diester, d is the closest edge-to-edge distance between cofactor atoms. 

Recent advances in measuring the kinetics of the various electron-transfer steps in this system have been achieved by use of flash photolysis of ruthenated derivatives of cytochrome c (Ru-Cc) (17-19). In these studies [Ru(bpy)3]2+ is covalently bound to a surface residue at a site that does not interfere with the docking of cytochrome c to cytochrome c oxidase. Solutions are then prepared containing both Ru-Cc and cytochrome c oxidase, and the two proteins associate to form a 1 1 complex. Flash photolysis of the solution leads directly to the excitation of the RuII(bpy)3 site, which then reduces heme c very rapidly. This method thus provides a convenient means to observe the subsequent intracomplex electron transfer from heme c to cytochrome c oxidase and further stages in the process. 

Increasing attention is being given to the reactivity of ruthenium species which show unusual behavior compared with their Co analogs. Aspects of current interest are mixed valence states, ruthenated proteins to probe electron transfer in them (Chap. 5) and the photochemistry and photophysics of Ru(II) polypyridine eomplexes. 

It is possible to introduce new redox groups into proteins using chemical and/ or biosynthetic methods. Attachment of mthenium to specific histidine residues on various electron transfer proteins has provided a method for the characterization of intramolecular electron transfer processes in proteins (77, 78). Studies of intramolecular electron transfer in ruthenated proteins are described in more detail in Section IV. 

Two intramolecular electron transfer systems of current interest, ruthenated proteins and photosynthetic reaction centers, are now discussed. These examples serve to illustrate the types of experimental approaches and information that may be obtained concerning the details of electron transfer processes in biological systems. 

Covalent attachment of ruthenium ions to proteins (ruthenation) provides a powerful approach for introducing a second redox active group into an electron transfer protein (77, 78). Under suitable conditions, intramolecular electron transfer may be monitored between Ru and the intrinsic redox group. Ruthenated proteins display a number of advantages for the study of intramolecular electron transfer reactions (77, 78) ... 

Selectively modified and characterized ruthenated proteins have been prepared for cytochrome c (141-144), cytochrome c-551 (145), azurin (146, 147), plastocyanin (147), HiPlP (/4S), and myoglobin (77, 149-154). Intramolecular electron transfer rates, k, measured in these systems are listed in Table 111 along... 

Rate constants of the individual electron transfer reactions are listed in Table IV, along with estimates of the driving force and the separation distance between cofactors. The rates of the first two steps (D — d>A and a Qa) are particularly impressive relative to the rates observed for comparable distances (10 A) and driving forces (—0.5 V) in ruthenated proteins. This suggests that more favorable nuclear and/or electronic factors have been achieved for the electron transfer reactions in the RC. 

Marcus theory predicts that the nuclear factor in the electron transfer rate expression will be maximal when - AG° = X. Under these conditions, the electron transfer process will be temperature independent. The first two electron transfer steps in the RC approximately exhibit this behavior. The rate of the D 4>a step actually increases slightly (by a factor of 2-4) as the temperature decreases from 300 to 8 K (180). Consequently, X may be estimated to be in the range 0.3 to 0.5 V (—7-10 kcal/mol) from the A ° values for these two steps (Table IV). These values are approximately 4 times smaller than those observed for ruthenated proteins discussed previously. Sequestration of the redox groups in a membrane-bound protein complex, away from aqueous solution, may serve to decrease the value of X by minimizing the reorganization energy of a highly polar solvent. 

The specific electron transfer characteristics of ruthenated proteins and photosynthetic RCs reflect relevant features of biological electron transfer systems in general ... 

For similar driving forces, a single rate-distance expression does not necessarily describe the behavior of different electron transfer systems. When all other factors are kept approximately constant, an exponential dependence of rate on distance is observed in ruthenated myoglobins. The nature of the intervening medium between donor and acceptor groups can significantly influence the rates of electron transfer observed in different systems, however. The presence of appropriate cofactors, amino acid side chains, and phytyl or isoprenoid tails between donors and acceptors appears to contribute to a substantial acceleration of the rate of electron transfer in RCs, relative to ruthenated proteins. 

Figure 9.21 Distance-dependence of rate constants for electron transfer in some protein systems. This plot shows the distance-dependence of the rate constant (optimised) for electron transfer between donor and acceptor atoms A and B on the distance between them, in protein systems of two types, (a) reaction centres in photosynthetic bacteria (o, ) and (b) ruthenated proteins (A, A). Diagram from Ref. [35]. See text.

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