Abstract

The triplet state of the primary donor in mutant reaction centers ofRhodobacter sphaeroides, in which amino acids near the primary donor were substituted, was investigated by absorbance-detected magnetic resonance. (ADMR). The mutations are associated with the substitution of leucines at site L131 and M160 near the bacteriochlorophyll halves of the primary donor by histidines, resulting in the formation of a hydrogen bond with the carbonyl group at ring V of either bacteriochlorophyll halves of the primary donor. For both mutant reaction centers the zero-field splitting parameters were slightly changed compared to native reaction centers, indicating that the dimer-halves of the primary donor in the latter are coupled in the triplet state: In addition, results from hole burning ADMR experiments on the mutants were compared with those forRhodobacter sphaeroides R26. The linewidths of the holes burnt in the zero-field transitions were similar for the mutant reaction centers involving the mutation at site L131 and for reaction centers ofRhodobacter sphaeroides R26; a somewhat larger linewidth was experiments on14N- and15N-containing reaction centers ofRhodobacter sphaeroides R26 showed that at high microwave power the holewidths of14N-reaction centers are determined by the quadrupole lines, but that at low microwave power the holewidths are mainly determined by unresolved hyperfine interactions with the protons. From the similarity in the holewidths for all reaction centers, we therefore conclude that the hyperfine interactions between the protons and the triplet spin, and thus the electronic composition of the triplet state, are similar for all reaction centers studied. The slight differences in the holewidths of the zero-field transitions and in the zero-field splitting parameters of the triplet state of the primary donor, and the differences previously observed for the interaction between the primary donor and neighboring bacteriochlorophylls (Vrieze J., Williams J.C., Allen J.P., Hoff A.J.: Biochim. Biophys. Acta1276, 221–228 (1996)), are attributed to small changes in charge-transfer contributions to the triplet state.

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