MUTATIONAL ANALYSIS OF PHOTOSYSTEM-I POLYPEPTIDES IN THE CYANOBACTERIUM SYNECHOCYSTIS SP, PCC-6803 - TARGETED INACTIVATION OF PSAI REVEALS THE FUNCTION OF PSAI IN THE STRUCTURAL ORGANIZATION OF PSAL

We cloned, characterized, and inactivated the psaI gene encoding a 4-k Da hydrophobic subunit of photosystem I from the cyanobacterium Synech ocystis sp. PCC 6803, The peal gene is located 90 base pairs downstrea m from psaL, and is transcribed on 0,94- and 0,SB-kilobase transcripts , To identify the function of psaI, we generated a cyanobacterial stra in in which psaI has been interrupted by a gene for chloramphenicol re sistance. The wild-type and the mutant cells showed comparable rates o f photoautotrophic growth at 25 degrees C, However, the mutant cells g rew slower and contained less chlorophyll than the wild-type cells, wh en grown at 40 degrees C. The PsaI-less membranes from cells grown at either temperature showed a small decrease in NADP(+) photoreduction r ate when compared to the wild-type membranes. Inactivation of psaI led to an 80% decrease in the PsaL level in the photosynthetic membranes and to a complete loss of PsaL in the purified photosystem I preparati ons, but had little effect on the accumulation of other photosystem I subunits, Upon solubilization with nonionic detergents, photosystem I trimers could be obtained from the wild-type, but not from the PsaI-le ss membranes, The PsaI-less photosystem I monomers did not contain det ectable levels of PsaL. Therefore, a structural interaction between Ps aL and PsaI may stabilize the association of PsaL with the photosystem I core. PsaL in the mild-type and PsaI-less membranes showed equal re sistance to removal by chaotropic agents. However, PsaL in the PsaI-le ss strain exhibited an increased susceptibility to proteolysis. From t hese data, we conclude that PsaI has a crucial role in aiding normal s tructural organization of PsaL within the photosystem I complex and th e absence of PsaI alters PsaL organization, leading to a small, but ph ysiologically significant, defect in photosystem I function.