Reduced turnover of the D1 polypeptide and photoactivation of electron transfer in novel herbicide resistant mutants of Synechocystis sp. PCC 6803.
Two missense mutants, A263P and S264P, and a deletion mutant des-Ala263, Ser264, have been constructed in the D1 protein of the cyanobacterium Synechocystis sp PCC 6803. All were expected to induce a significant conformational change in the QB-binding region of photosystem II (PSII). Although the des-Ala263, Ser264-D1 mutant accumulated some D1 protein in the thylakoid membrane it was unable to grow photoautotrophically or evolve oxygen. Thermoluminescence and chlorophyll fluorescence studies confirmed that this deletion mutant did not show any functional PSII activity. In contrast, [S264P]D1 was able to grow photoautotrophically and give light-saturated rates of oxygen evolution at 60% of the rate of the wild-type control strain, TC31. The A263P missense mutant was also able to evolve oxygen at 50% of TC31 rates although it did not readily grow photoautotrophically. Thermoluminescence, flash oxygen yield and chlorophyll fluorescence measurements indicated that in both missense mutants electron transfer from QA to QB was significantly impaired in dark adapted cells. However, QA to QB electron transfer could be photoactivated in the mutants by background illumination. Both the A263P and S264P mutants also showed an increase in resistance to the s-triazine family of herbicides although this feature did not hold for the phenolic herbicide, ioxynil. Of particular interest was that the two missense mutants, especially S264P, possessed a slower rate of turnover of the D1 protein compared with TC31 and in vivo contained detectable levels of a 41-kDa adduct consisting of D1 and the alpha subunit of cytochrome b559. When protein synthesis was blocked by the addition of lincomycin, D1 degradation was again slower in S264P than TC31. The results are discussed in terms of structural changes in the QB-binding region which affect herbicide and plastoquinone binding and perturb the normal regulatory factors that control the degradation of the D1 protein and its synchronisation with the synthesis of a replacement D1 protein.
Dalla Chiesa
M
M
Department of Biochemistry, Imperial College of Science, Technology and Medicine, London, UK.
Friso
G
G
Deák
Z
Z
Vass
I
I
Barber
J
J
Nixon
P J
PJ
D001665:Binding Sites; D003001:Cloning, Molecular; D000458:Cyanobacteria / Q000254:growth & development / Q000378:metabolism*; D004351:Drug Resistance; D004579:Electron Transport; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005453:Fluorescence; D006540:Herbicides / Q000494:pharmacology*; D007462:Iodobenzenes / Q000494:pharmacology; D008027:Light; D045342:Light-Harvesting Protein Complexes; D008034:Lincomycin / Q000494:pharmacology; D008163:Luminescent Measurements; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009570:Nitriles / Q000494:pharmacology; D010100:Oxygen / Q000378:metabolism; D010788:Photosynthesis; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D045332:Photosystem II Protein Complex; D013696:Temperature; D014227:Triazines / Q000494:pharmacology
D006540:Herbicides; D007462:Iodobenzenes; D045342:Light-Harvesting Protein Complexes; D008565:Membrane Proteins; D009570:Nitriles; D045322:Photosynthetic Reaction Center Complex Proteins; D045332:Photosystem II Protein Complex; D014227:Triazines; C008161:ioxynil; D008034:Lincomycin; D010100:Oxygen