BINDING AND INTERACTION OF THE PRIMARY AND THE SECONDARY-ELECTRON ACCEPTOR QUINONES IN BACTERIAL PHOTOSYNTHESIS - AN INFRARED SPECTROELECTROCHEMICAL STUDY OF RHODOBACTER-SPHAEROIDES REACTION CENTERS

The vibrational modes of the primary and secondary electron acceptors Q(A) and Q(B), their semiquinone anions and their respective protein e nvironment in Rhodobacter sphaeroides reaction centers have been chara cterized using combinations of electrochemically-induced and light-ind uced Fourier-transform infrared (IR) difference spectroscopy. Q(A)-/Q( A) and Q(B)-/Q(B) IR difference spectra without contributions of other cofactors were generated by three different methods: (1) electrochemi cally, by reduction of Q(A) to Q(A)-; (2) photochemically, with flash- induced formation of P+Q(A)- or P+Q(B)-, rereduction of P+ by cytochro me (cyt) c2, and electrochemical rereduction of cyt c2; (3) photochemi cally and electrochemically, by subtraction of redox-induced P+/P diff erence spectra from light-induced P+Q(A)-/PQ(A) and P+Q(B)-/PQ(B) diff erence spectra. Although Q(A)- was generated by completely different m ethods, and in one case (3) a charge-separated state is involved, almo st identical Q(A)-/Q(A) and only slightly different Q(B)-/Q(B) differe nce spectra have been obtained. Bands at 1630 cm-1 and 1640 cm-1 are p roposed as candidates for the C=O modes of Q(A) and Q(B), respectively . The C-O modes of Q(A)- and Q(B)- are assigned to bands at 1462 cm-1 and 1478-88 cm-1, respectively. Difference bands at 1668 cm-1 and 1652 cm-1 in Q(A)-/Q(A) difference spectra are more likely to arise from a mide-I modes or side chain vibrations of amino acids to which Q(A) is hydrogen-bonded. A number of difference bands between 1520 cm-1 and 15 60 cm-1 possibly arise from amide-II vibrations and aromatic amino-aci d side chain residues in the vicinity Of Q(A) and Q(B). A differential feature at 1734 cm-1/1726 cm-1 in Q(A-)/Q(A) difference spectra proba bly arises from changes in the protonation state or environment of dis tant carboxyl groups. An alternative explanation in terms of changes i n the environment of the 10a ester C=O group of bacteriopheophytin L, however, cannot be excluded. Bands between 1724 cm-1 and 1740 cm-1 in Q(B)-/Q(B) difference spectra are tentatively assigned to a protonatio n of ASP L213 and/or a change in the environment of GLU L212, both bei ng located in the vicinity Of Q(B) and involved in the proton transfer to Q(B) (Okamura, M.Y. and Feher, G. (1992) Annu. Rev. Biochem. 61, 8 61-896).