Isolation and characterization of reaction centre-antenna conjugates from a range of bacteriochlorophyll a-containing species of purple bacteria

Dawkins, Deborah J (1988) Isolation and characterization of reaction centre-antenna conjugates from a range of bacteriochlorophyll a-containing species of purple bacteria. PhD thesis, University of Glasgow.

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The photosynthetic apparatus of purple bacteria is made up of two major components; a well-characterized reaction centre, and light-harvesting antenna complexes. The latter complexes are responsible for absorbing incident radiation and transferring it to the reaction centre where it is utilized in the primary processes of photosynthesis. The antenna complex consists typically of a small number of bacteriochlorophyll a (2-3) and carotenoid molecules (1 or 2) bound noncovalently to two low molecular weight (5-7kd) hydrophobic polypeptides [Thornber et al, 1983]. Two types of antenna complex, which are differentiated with respect to their NIR absorption maxima, were recognized in the early studies of Aagaard and Sistrom (1972). The first type designated the B875 (B880) antenna complex, appears to be intimately associated with the reaction centre, whereas the second type, the B800-850 complex, having no apparent fixed relationship with the reaction centre, is present in variable amounts depending upon the prevailing environmental conditions. The relationship existing between the reaction centre and B875 (B880) complex is emphatically illustrated in the photosynthetic membranes of bacteriochlorophyll b-containing species of purple bacteria, in particular of Rhodopseudomonas viridis and Ectothiorhodospira halochloris. Examination of these membranes, using electron microscopy and image-processing techniques, shows the membranes to consist of an array of well-defined, identical structures, consisting of a reaction centre surrounded by, and complexed with, the B875-type (or B1012 in Rps. viridis) of antenna complex. To date, a number of studies on bacteriochlorophyll a-containing species, including developmental, structural, immunofractionation, solubilization and functional analyses, have provided positive evidence supporting the existence of a fixed stoichiometric relationship between the reaction centre and the B875 (B880) antenna complex. However, whether this reflects the existence of a fixed stoichiometric core structure, similar to that observed in BChl b-containing species, still remains to be seen. The method described by Firsow and Drews (1977) for the fractionation of detergent solubilized membranes has been modified in this study for the isolation of spectrally-pure RC-B875 and B800-850 antenna complexes from a range of BChl a-containing species of purple photosynthetic bacteria. Solubilization conditions were optimized for each species examined. The ratio of bacteriochlorophyll a present per reaction centre in the isolated RC-B875 conjugates was then determined from their respective absorption and oxidation-reduction difference spectra. For each species the ratio of RC:BChl a was constant irrespective of the light intensity at which the cells were grown. In making comparisons between species a degree of variation was apparent. Subsequent examination of the effects of additional, or alternative, detergents on this ratio value however, suggests that the variation may be a consequence of the experimental procedures adopted rather than the result of true species variation. The fluorescence induction kinetics of these isolated complexes were also examined in order to determine their functional state. The successful detection of variable fluorescence which depends upon the redox state of the reaction centre, suggests that the light-harvesting complexes are still functionally connected to the reaction centre in the isolated complexes. The polypeptide composition was confirmed by SDS-polyacrylamide gel electrophoresis. Finally in a quest to determine the organizational structure of the RC-antenna conjugate crystallization attempts have been made. It appears from our results that all purple bacteria contain a common antenna-reaction centre "core" structure that forms the basis of their photosynthetic unit.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Keywords: Microbiology
Date of Award: 1988
Depositing User: Enlighten Team
Unique ID: glathesis:1988-76693
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 19 Nov 2019 13:53
Last Modified: 19 Nov 2019 13:53

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