To obtain information on the structural and functional role of highly conserved amino acid residues in the B870 and light-harvesting polypeptides of Rhodobucter cupsulutus, site-directed mutagenesis was performed. 18 mutants with single amino acid substitutions at nine different positions in the B870 antenna polypeptides were prepared in a B800-850-lacking strain. The characterization of the resulting phenotypes was based on a quantification of the core-complex elements (reaction center, light-harvesting polypeptides, bacteriochlorophyll u and carotenoid) and the core-complex spectral characteristics (absorption maximum, absorption coefficient and fluorescence intensity). These data generally showed that strong structural effects were caused by the amino acid substitutions. Thus, the three tryptophan exchanges at the position cr8 resulted in either the absence of a core complex (aTrp8-+Leu), the absence of the core antenna (aTrpS+Ala) or a reduction in the carotenoid content (aTrp8-tTyr). Likewise, the mutants aProl3Gly (i.e. aProl3-+Gly), PGlylOVal and aPhe23Ala demonstrated an abnormal protein/pigment ratio in the core antenna, while a drastically reduced antenna size resulted from the amino acid exchange PArg45Asp. In contrast to the structural effects, the absorption maxima and the fluorescence intensities of the mutant antennae differed only slightly from the wild type. The strongest blue shift of the bacteriochlorophyll u (8-11 nm) was induced by substitutions of the Trp at position a43 (aTrp43-+Ala, Leu or Tyr). Contrary to the other spectral effects, the absorption coefficient of bacteriochlorophyll u was strongly influenced by the amino acid substitutions and varied by 1.6-times less (pArg45Asp) and 1.3-times greater (aPhe25Ala) than normal. The antenna-free mutant, aTrpSAla, yielded a high rate of B800-850 revertants during phototrophic growth, indicating a direct energy transfer from the B800-850 antenna to the reaction center in these strains. Although conditions for growth were generally observed to influence phenotypic expression, the structural as well as spectral effects were demonstrated to differ to the greatest extent between chemotrophically grown and phototrophically grown cells.Rhodobacter capsulatus is a Gram-negative, photosynthetic bacterium and belongs to the group of non-sulfur purple bacteria [l]. The photosynthetic system of Rb. cupsulutus consists of two functional units; the light-harvesting antenna which absorbs light energy and transfers it to the reaction center (RC) and the RC which applies this energy to produce a proton gradient across the membrane. The antenna is an intramembranous protein-pigment complex. Two antenna complexes, the B870 and B800-850 complexes, have been structurally and spectroscopically distinguished in Rb. cupsulutus, and these are subsequently named according to their absorption peaks in the near-infrared region (Q, band). The B870 antenna forms a fixed-size aggregate with the RC, called the core complex. This complex is surrounded by the B800-850 comple...
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