Deletion of the PEST-like region of photosystem two modifies the QB-binding pocket but does not prevent rapid turnover of D1.
The rapid turn-over of the D1 polypeptide of the photosystem two complex has been suggested to be due to the presence of a "PEST"-like sequence located between putative transmembrane helices IV and V of D1 (Greenberg, B. M., Gaba, V., Mattoo, A. K. and Edelman, M. (1987) EMBO J. 6, 2865-2869). We have tested this hypothesis by constructing a deletion mutant (delta 226-233) of the cyanobacterium Synechocystis sp. PCC 6803 in which residues 226-233 of the D1 polypeptide, containing the PEST-like sequence, have been removed. The resulting mutant, delta PEST, is able to grow photoautotrophically and give light-saturated rates of oxygen at wild type levels. However electron transfer on the acceptor side of the complex is perturbed. Analysis of cells by thermoluminescence and by monitoring the decay in quantum yield of variable fluorescence following saturating flash excitation indicates that Q-B, but not Q-A, is destabilized in this mutant. Electron transfer on the donor side of photosystem two remains largely unchanged in the mutant. Turnover of the D1 polypeptide as examined by pulse-chase experiments using [35S]methionine was enhanced in the delta PEST mutant compared to strain TC31 which is the wild type control. We conclude that the PEST sequence is not absolutely required for turnover of the D1 polypeptide in vivo although deletion of residues 226-233 does have an effect on the redox equilibrium between QA and QB.
Nixon
P J
PJ
Wolfson Laboratories, Biochemistry Department, Imperial College of Science, Technology and Medicine, London, United Kingdom.
Komenda
J
J
Barber
J
J
Deak
Z
Z
Vass
I
I
Diner
B A
BA
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D000458:Cyanobacteria / Q000378:metabolism*; D007700:Kinetics; D008027:Light; D008163:Luminescent Measurements; D046911:Macromolecular Substances; D008715:Methionine / Q000378:metabolism; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D045322:Photosynthetic Reaction Center Complex Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D045332:Photosystem II Protein Complex; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D013462:Sulfur Radioisotopes
D046911:Macromolecular Substances; D045322:Photosynthetic Reaction Center Complex Proteins; D045332:Photosystem II Protein Complex; D011994:Recombinant Proteins; D013462:Sulfur Radioisotopes; D008715:Methionine