Beschreibung:
The photosynthetic machinery of the thylakoid membranes comprises about 100 protein complexes. Among them the light harvesting complex protein family forms a large subgroup, their large number bearing from a large number of nuclear-encoded genes, as well as posttranslational modifications. In this study 2-DE separation of thylakoid membranes from Chlamydomonas reinhardtii has been established as a tool for proteome mapping and functional analysis of light harvesting proteins, and dissection of mutant phenotypes affected in the expression and modification of thylakoid proteins. A mutant screen has been applied to study the assembly of light harvesting proteins. In this screen a Chl b-deficient mutant was obtained that was characterised biochemically and physiologically. PAGE analyses and oxygen uptake assays implied that LHC proteins are reduced and detached from the photosystem. In the Chl b-deficient mutant cbs3 similar findings were obtained. Whereas effects on LHC proteins could be caused by Chl b-deficiency in cbs3, Chl b-deficiency in S2 is implied to be a consequence of LHC destabilisation, although the impact of regulation mechanisms in the context of pigment biosynthesis cannot be ruled out so far. Further pigment analysis looking at different growth conditions and measurement of npq could be valuable tools, especially if cbs3 is included in the analysis. Unfortunately although highly elaborate state of the art skills were applied it was not possible to perform genetic crosses with the mutant S2 in order to correlate the introduction of the ble-gene with the phenotype of the mutant. After the mutant has been characterised biochemically in this study it would now be worthwhile to identify the affected DNA-segment by plasmid rescue.