The ras protein family: evolutionary tree and role of conserved amino acids.

Preresonance Raman and resonance Raman spectra of the primary donor (P) from reaction centers of the Rhodobacter (Rb. ) sphaeroides R26 carotenoidless strain in the P and P+ states, respectively, were obtained at room temperature with 1064-nm excitation and a Fourier transform spectrometer. These spectra clearly indicate that the chromophore modes are observable over those of the protein with no signs of interference below 1800 cm-*. The chromophore modes are dominated by those of the bacteriochlorophylls (BChl a) , and it is estimated that, in the P state, ca. 65% of the Raman intensity of the BChl a modes arises from the primary donor. This permits the direct observation of a vibrational spectrum of the primary donor at preresonance with the excitonic 865-nm band. The Raman spectrum of oxidized reaction centers in the presence of ferricyanide clearly exhibits bands arising from a BChl a+ species. The magnitude of the frequency shift of a keto carbonyl of neutral P from 1691 to 17 17 cm-' upon P+ formation strongly suggests that one BChl molecule in P+ carries nearly the full +1 charge. Our results indicate that the unpaired electron in P+ does not share a molecular orbital common to the two components of the dimer on the time scale of the resonance Raman effect (ca. s). x e primary events in bacterial photosynthesis occur in membrane-bound proteins known as reaction centers (RCs).' The isolated RC consists of six bacteriochlorin pigments (four bacteriochlorophyll a and two bacteriopheophytin a molecules), two quinones, one non-heme iron, one carotenoid molecule, and approximately 850 amino acid residues contained in three. polypeptide subunits named L, M, and H. Within the RC, electron transfer originates from the primary donor P, which consists of a pair of bacteriochlorophyll (BChl) molecules in mutual excitonic interaction. Although the X-ray crystallographic structures of the RC from Rhodopseudomonas (Rps.) viridis (Deisenhofer & Michel, 1989) and Rhodobacter (Rb.) sphaeroides (Allen et al., 1987a,b; Chang et al., 1986; T i d e et ai., 1988) are resolved, the understanding of charge separation and stabilization requires a thorough characterization of the physicochemical properties of P and its cation radical, P+. The absorption spectrum of bacterial reaction centers exhibits a broad band in the near-infrared that corresponds to the first excited singlet state of the primary donor pair, 'P. For bacteriochlorophyll a (BChl a ) containing RCs, such as Rb. sphaeroides, this band appears at ca. 870 nm. The characterization of this band in an attempt to explain the asymmetric functioning of the RC has been the subject of recent intensive work [for a review, see Friesner and Won (1989)]. When P undergoes one-electron chemical or phoT.A.M. gratefully acknowledges fellowships from NATO/NSERC Abbreviations: RR, resonance Raman; NIR, near-infrared; FT, Fourier transform; RC, reaction center; Rb., Rhodobacter; Rps., Rhodopseudomonas; Rsp., Rhodospirillum; BChl, bacteriochlorophyll; BPhe, bacteriopheophytin; EPR, electron paramagnetic resonance; THF, tet(Canada) and EMBO. *Author to whom correspondence should be addressed. *CE Saclay. * Universitit Essen. rahydrofuran. 0006-2960/9 1 /0430-4648$02.50/0 0 199 1 American Chemical Society