bacteriochlorophylls and geranylgeraniol

The Fenna-Matthews-Olson light-harvesting antenna (FMO) protein has been a model system for understanding pigment-protein interactions in the energy transfer process in photosynthesis. All previous studies have utilized wild-type FMO proteins from several species. Here we report the purification and characterization of the first FMO protein variant generated via replacement of the esterifying alcohol at the C-17 propionate residue of bacteriochlorophyll (BChl) a, phytol, with geranylgeraniol, which possesses three more double bonds. The FMO protein still assembles with the modified pigment, but both the whole cell absorption and the biochemical purification indicate that the mutant cells contain a much less mature FMO protein. The gene expression was checked using qRT-PCR, and none of the genes encoding BChl a-binding proteins are strongly regulated at the transcriptional level. The smaller amount of the FMO protein in the mutant cell is probably due to the degradation of the apo-FMO protein at different stages after it does not bind the normal pigment. The absorption, fluorescence, and CD spectra of the purified FMO variant protein are similar to those of the wild-type FMO protein except the conformations of most pigments are more heterogeneous, which broadens the spectral bands. Interestingly, the lowest-energy pigment binding site seems to be unchanged and is the only peak that can be well resolved in 77 K absorption spectra. The excited-state lifetime of the variant FMO protein is unchanged from that of the wild type and shows a temperature-dependent modulation similar to that of the wild type. The variant FMO protein is less thermally stable than the wild type. The assembly of the FMO protein and also the implications of the decreased FMO/chlorosome stoichiometry are discussed in terms of the topology of these two antennas on the cytoplasmic membrane.

Topics: Bacterial Proteins; Bacteriochlorophyll A; Chlorobi; Diterpenes; Energy Transfer; Esterification; Gene Expression Profiling; Gene Expression Regulation, Bacterial; Genetic Variation; Light-Harvesting Protein Complexes; Photosynthesis; Phytol; Protein Stability

The absorption and fluorescence properties of chlorosomes of the filamentous anoxygenic phototrophic bacterium Chloronema sp. strain UdG9001 were analyzed. The chlorosome antenna of Chloronema consists of bacteriochlorophyll (BChl) d and BChl c together with gamma-carotene as the main carotenoid. HPLC analysis combined with APCI LC-MS/MS showed that the chlorosomal BChls comprise a highly diverse array of homologues that differ in both the degree of alkylation of the macrocycle at C-8 and/or C-12 and the alcohol moiety esterified to the propionic acid group at C-17. BChl c and BChl d from Chloronema were mainly esterified with geranylgeraniol (33% of the total), heptadecanol (24%), octadecenol (19%), octadecanol (14%), and hexadecenol (9%). Despite this pigment heterogeneity, fluorescence emission of the chlorosomes showed a single peak centered at 765 nm upon excitation at wavelengths ranging from 710 to 740 nm. This single emission, assigned to BChl c, indicates an energy transfer from BChl d to BChl c within the same chlorosome. Likewise, incubation of chlorosomes under reducing conditions caused a weak increase in fluorescence emission, which indicates a small redox-dependent fluorescence. Finally, protein analysis of Chloronema chlorosomes using SDS-PAGE and MALDI-TOF-MS revealed the presence of a chlorosomal polypeptide with a molecular mass of 5.7 kDa, resembling the CsmA protein found in Chloroflexus aurantiacus and Chlorobium tepidum chlorosomes. Several minor polypeptides were also detected but not identified. These results indicate that, compared with other members of filamentous anoxygenic phototrophic bacteria and green sulfur bacteria, Chloronema possesses an antenna system with novel features that may be of interest for further investigations.

Topics: Bacterial Proteins; Bacteriochlorophylls; Carotenoids; Chloroflexi; Chromatography, High Pressure Liquid; Diterpenes; Light-Harvesting Protein Complexes; Spectrometry, Fluorescence; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Spectrophotometry; Spectrum Analysis

A gene from the cyanobacterium Synechocystis sp. PCC 6803 has been cloned and sequenced, and subsequently used to partially complement a bchP mutant of the purple photosynthetic bacterium Rhodobacter sphaeroides. This mutant is blocked in the terminal hydrogenation steps of bchla biosynthesis and possesses only bchl esterified with geranylgeraniol. It also has a reduced cellular level of the light-harvesting LH2 complex, and the 850 nm absorbance maximum of LH2 is red-shifted by approximately 6 nm. Upon heterologous expression of the Synechocystis bchP homologue, not only are hydrogenated forms of bchlaGG detectable, but the level of LH2 is increased and the red-shift reversed by several nm. We conclude that this gene, which we term chlP, encodes the enzyme catalysing the stepwise hydrogenation of geranylgeraniol to phytol during chla biosynthesis.

Topics: Amino Acid Sequence; Bacterial Proteins; Bacteriochlorophylls; Base Sequence; Cloning, Molecular; Cyanobacteria; Diterpenes; Genetic Complementation Test; Hydrogenation; Light-Harvesting Protein Complexes; Molecular Sequence Data; Phenotype; Photosynthetic Reaction Center Complex Proteins; Phytol; Rhodobacter sphaeroides; Sequence Analysis, DNA; Sequence Homology, Amino Acid; Spectrophotometry

Four chemically different bacteriochlorophylls (Bchls) a esterified with geranylgeraniol, dihydrogeranylgeraniol, tetrahydrogeranylgeraniol, and phytol have been detected by high-pressure liquid chromatography in cell extracts from Rhodopseudomonas sphaeroides and Chromatium vinosum. Bchl a containing phytol is the principal component, and the other three Bchls a comprise about 4% of the total Bchls a in stationary-phase cells of R. sphaeroides and C. vinosum. The high levels of the minor pigments occur in the beginning of Bchl a phytol formation, indicating that they are not degradation products, but intermediates of Bchl a phytol formation.

Topics: Bacteriochlorophylls; Chlorophyll; Chromatium; Chromatography, High Pressure Liquid; Diterpenes; Esterification; Phytol; Rhodobacter sphaeroides