In Rhodobacter sphaeroides Reaction Centers, Mutation of Proline L209 to Aromatic Residues in the Vicinity of a Water Channel Alters the Dynamic Coupling between Electron and Proton Transfer Pr
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- 作者：Julia Tandori ; Pierre Sebban ; Hartmut Michel ; and Laura Baciou
- 刊名：Biochemistry
- 出版年：1999
- 出版时间：October 5, 1999
- 年：1999
- 卷：38
- 期：40
- 页码：13179 - 13187
- 全文大小：111K
- 年卷期：v.38,no.40(October 5, 1999)
- ISSN：1520-4995
文摘
The X-ray crystallographic structure of the photosynthetic reaction center from Rhodobactersphaeroides obtained at high resolution has revealed a number of internal water molecules (Ermler, U.,Fritzsch, G., Buchanan, S. K., and Michel, H. (1994) Structure 2, 925-936; Stowell, M. H. B., McPhillips,T. M., Rees, D. C., Soltis, S. M., Abresch, E., and Feher, G. (1997) Science 276, 812-816). Some ofthem are organized into distinct hydrogen-bonded water chains that connect QB (the terminal quinoneelectron acceptor of the reaction center) to the aqueous phase. To investigate the role of the water chainsin the proton conduction process, proline L209, located immediately adjacent to a water chain, was mutatedto the following residues: F, Y, W, E, and T. We have first analyzed the effects of the mutations on thekinetic and thermodynamic properties of the rate constants of the second electron transfer (kAB(2)) and ofthe coupled proton uptake (kH+) at the second flash. In all aromatic mutants, kAB(2) and kH+ are notablyand concomitantly decreased compared to the wild-type, while no effect is observed in the other mutants.The temperature dependence of these rates shows activation energy values (
H
) similar for the protonand electron-transfer processes in the wild-type and in most of the mutants, except for the L209PW andL209PF mutants. The analysis of the enthalpy factors related to the electron and proton-transfer processesin the L209PF and the L209PW mutants allows to distinguish the respective effects of the mutations forboth transfer reactions. It is noteworthy that in the aromatic mutants a substantial increase of the freeenergies of activation is observed (GL209PY < GL209PF < GL209PW) for both proton and electron-transfer reactions, while in the other mutants, G is not affected. The salt concentration dependence ofkAB(2) shows, in the L209PF and L209PW mutants, a higher screening of the protein surface potentialexperienced by QB. Our data suggest that residues F and W in position L209 increase the polarizabilityof the internal water molecules and polar residues by altering the organization of the hydrogen-bondnetwork. We have also analyzed the rates of the first electron-transfer reaction (kAB(1)), in the 100 
stime domain. These kinetics have previously been shown to reflect protein relaxation events possiblyincluding proton uptake events (Tiede, D. M., Vazquez, J., Cordova, J., and Marone, P. M. (1996)Biochemistry 35, 10763-10775). Interestingly, in the L209PF and L209PW mutants, kAB(1) is notablydecreased in comparison to the wild type and the other mutants, in a similar way as kAB(2) and kH+. Ourdata imply that the dynamic organization of this web is tightly coupled to the electron transfer processthat is kinetically limited by protonation events and/or conformational rearrangements within the protein.
H) similar for the protonand electron-transfer processes in the wild-type and in most of the mutants, except for the L209PW andL209PF mutants. The analysis of the enthalpy factors related to the electron and proton-transfer processesin the L209PF and the L209PW mutants allows to distinguish the respective effects of the mutations forboth transfer reactions. It is noteworthy that in the aromatic mutants a substantial increase of the freeenergies of activation is observed (GL209PY < GL209PF < GL209PW) for both proton and electron-transfer reactions, while in the other mutants, G is not affected. The salt concentration dependence ofkAB(2) shows, in the L209PF and L209PW mutants, a higher screening of the protein surface potentialexperienced by QB. Our data suggest that residues F and W in position L209 increase the polarizabilityof the internal water molecules and polar residues by altering the organization of the hydrogen-bondnetwork. We have also analyzed the rates of the first electron-transfer reaction (kAB(1)), in the 100 stime domain. These kinetics have previously been shown to reflect protein relaxation events possiblyincluding proton uptake events (Tiede, D. M., Vazquez, J., Cordova, J., and Marone, P. M. (1996)Biochemistry 35, 10763-10775). Interestingly, in the L209PF and L209PW mutants, kAB(1) is notablydecreased in comparison to the wild type and the other mutants, in a similar way as kAB(2) and kH+. Ourdata imply that the dynamic organization of this web is tightly coupled to the electron transfer processthat is kinetically limited by protonation events and/or conformational rearrangements within the protein.