Hydrogen Bonding and Circular Dichroism of Bacteriochlorophylls in the Rhodobacter
capsulatus Light-harvesting 2 Complex Altered by Combinatorial Mutagenesis
Hu Q., Sturgis J.N., Robert B., Delagrave S., Youvan D.C., Niederman R.A.
Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, New
Jersey, 08855-1059, USA.
We have investigated the spectroscopic properties of two classes of light-harvesting 2
(LH2, B800-850) mutants of Rhodobacter capsulatus obtained by combinatorial
mutagenesis to the C-terminal half of the beta-apoprotein: a pseudoLH2 (pLH2) class, in
which the 800-nm absorption was normal but the 850-nm peak was blue-shifted by up to 14
nm, and the other a pseudoLH1 (pLH1) class, which lacked the 800-nm absorption band and
showed 850-nm absorption red-shifts of up to 30 nm. In several of the pLH1 antennae,
carotenoid depletion contributed to the phenotype, while in the pLH2 complexes there was
some carotenoid enrichment. A number of mutants from each class have also been
characterized by low-temperature absorption and fluorescence spectroscopy, resonance Raman
spectroscopy, and circular dichroism. In all of the mutants investigated, the B850
bacteriochlorophyll a binding site remained intact, conserving both the hydrogen bonding
environment of the chromophores and their conformation and liganding. In contrast, the
intensity of the CD spectra of pLH1 complexes was considerably reduced, relative to that
of wild-type or pLH2 complexes, consistent with alterations in the interactions between
pigments and in their relative orientation. Elevated fluorescence polarization over the
red wing of the B850 band in the pLH2 complexes indicated a reduction of exciton mobility
within the ring of BChl molecules. Possible structural alterations governing the spectral
properties of the different mutants are discussed.