SITE-DIRECTED MODIFICATION OF THE LIGANDS TO THE BACTERIOCHLOROPHYLLSOF THE LIGHT-HARVESTING LH1 AND LH2 COMPLEXES OF RHODOBACTER-SPHAEROIDES

The core light-harvesting LH1 complex of Rhodobacter sphaeroides consi sts of an assembly of membrane-spanning alpha and beta polypeptides, e ach of which binds one bacteriochlorophyll molecule. In this study we have used site-directed mutagenesis to demonstrate that the B880 bacte riochlorophyll binding site of LH1 shows a high degree of specificity for the residue that provides the ligand to the Bchl Mg2+ ion. alpha H is(0) (alpha H-0) was changed to Asn, Leu, and Tyr, and beta His(0) (b eta H-0) to Asn, Gln, Leu, and Tyr; the mutated genes were expressed t o yield both LH1-only and LH1 + RC strains, in two different carotenoi d backgrounds, None of the alpha H-0 mutations formed an LH1 complex e ither in isolation or with RCs. Of the mutations of beta H-0 those to Asn and Gln formed LH1 complexes but in the latter case the complex wa s very unstable and occurred at very low cellular levels. In a similar study, the alpha H-0 and beta H-0 residues of the LH2 complex were ch anged to Asn. However, no complex was formed in either case. FT Raman spectroscopy of the beta H0N mutant LH1 shows perturbation of the inte raction state of the keto carbonyl of one Bchl which sheds light on th e possible H-bond partners for these keto oxygens. These data directly address the B880 binding site of the core LH1 complex and show that i t is subtly different from the B850 binding site of the peripheral LH2 complex. A model of this binding site may be proposed from these resu lts.