Probing the B800 bacteriochlorophyll binding site of the accessory light-harvesting complex from Rhodobacter sphaeroides using site-directed mutants. I. Mutagenesis, effects on binding, function and electrochromic behaviour of its carotenoids

Abstract

The light-harvesting LH2 complex of Rhodobacter sphaeroides contains two amino acid residues, βHis21 and βArg29, which are conserved in all LH2 β-polypeptides of purple nonsulfur bacteria sequenced so far. These residues have been changed into serine and glutamic acid, respectively. Both mutations lead to severe changes in the spectroscopic characteristics of the antenna complex. Changing βArg29 into Glu results in a blue shift and a broadening of the B800 bacteriochlorophyll absorbance, suggesting a role of this residue in creating the binding pocket for B800 (see also Visschers et al. (1994) Biochim. Biophys Acta 1183, 483–490). Similar blue shifts, of approx. 6 nm, are also observed in the carotenoid absorbance peaks. This is accompanied by a large change in the electrochromic behaviour of the carotenoids, which suggests a major role of βArg29 in creating a local field near the responsive carotenoid. The second mutation, βHis21 to Ser, results in an inability to create a B800 domain. This mutation also causes changes in the carotenoid absorbance and electrochromic behaviour, suggesting a direct or indirect (via the bacteriochlorophyll B800 molecule) effect on the local dipole field of the sensitive carotenoid. Neither of the mutated complexes has lost the ability to bind carotenoids; in both complexes energy transfer from the carotenoids to B850 appears unaltered, indicating that all carotenoids can transfer energy directly to this bacteriochlorophyll, despite the loss of B800.