A HOLE-BURNING STUDY ON RHODOBACTER-SPHAEROIDES WITH ABSORBENCY-DETECTED MAGNETIC-RESONANCE (ADMR)

The triplet state of the primary donor in mutant reaction centers of R hodobacter sphaeroides, in which amino acids near the primary donor we re substituted was investigated by absorbance-detected magnetic resona nce (ADMR). The mutations are associated with the substitution of leuc ines 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 h alves of the primary donor. For both mutant reaction centers the zero- field splitting parameters were slightly changed compared to native re action centers, indicating that the dimer-halves of the primary donor in the latter are coupled in the triplet stale. In addition, results f rom hole burning ADMR experiments on the mutants were compared with th ose for Rhodobacter sphaeroides R26. The linewidths of the holes burnt in the zero-field transitions were similar for the mutant reaction ce nters involving the mutation at site L131 and for reaction centers of Rhodobacter sphaeroides R26; a somewhat larger linewidth was found for the reaction centers involving the mutation at site M160. Comparison of the results from experiments on N-14- and N-15-containing reaction centers of Rhodobacter sphaeroides R26 showed that at high microwave p ower the holewidths of N-14-reaction centers are determined by the qua drupole lines, but that at low microwave power the holewidths are main ly determined by unresolved hyperfine interactions with the protons. F rom the similarity in the holewidths for all reaction centers, we ther efore 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 differ ences 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 th e primary donor and neighboring bacteriochlorophylls (Vrieze J., Willi ams J.C., Alien J.P., Hoff A.J.: Biochim. Biophys. Acta 1276, 221-228 (1996)), are attributed to small changes in charge-transfer contributi ons to the triplet state.