title: 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.
abstract: We cloned, characterized, and inactivated the psaI gene encoding a 4-kDa hydrophobic subunit of photosystem I from the cyanobacterium Synechocystis sp. PCC 6803. The psaI gene is located 90 base pairs downstream from psaL, and is transcribed on 0.94- and 0.32-kilobase transcripts. To identify the function of PsaI, we generated a cyanobacterial strain in which psaI has been interrupted by a gene for chloramphenicol resistance. The wild-type and the mutant cells showed comparable rates of photoautotrophic growth at 25 degrees C. However, the mutant cells grew slower and contained less chlorophyll than the wild-type cells, when grown at 40 degrees C. The PsaI-less membranes from cells grown at either temperature showed a small decrease in NADP+ photoreduction rate 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 preparations, 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-less membranes. The PsaI-less photosystem I monomers did not contain detectable levels of PsaL. Therefore, a structural interaction between PsaL and PsaI may stabilize the association of PsaL with the photosystem I core. PsaL in the wild-type and PsaI-less membranes showed equal resistance to removal by chaotropic agents. However, PsaL in the PsaI-less strain exhibited an increased susceptibility to proteolysis. From these data, we conclude that PsaI has a crucial role in aiding normal structural organization of PsaL within the photosystem I complex and the absence of PsaI alters PsaL organization, leading to a small, but physiologically significant, defect in photosystem I function.
authors.affiliation: Division of Biology, Kansas State University, Manhattan 66506-4901, USA.
authors.forename: Q
authors.initials: Q
authors.lastname: Xu
authors.forename: D
authors.initials: D
authors.lastname: Hoppe
authors.forename: V P
authors.initials: VP
authors.lastname: Chitnis
authors.forename: W R
authors.initials: WR
authors.lastname: Odom
authors.forename: J A
authors.initials: JA
authors.lastname: Guikema
authors.forename: P R
authors.initials: PR
authors.lastname: Chitnis
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics*; D015152:Blotting, Northern; D002918:Chymotrypsin / Q000494:pharmacology; D003001:Cloning, Molecular; D003432:Cross-Linking Reagents; D000458:Cyanobacteria / Q000235:genetics* / Q000378:metabolism / Q000528:radiation effects; D004252:DNA Mutational Analysis; D015151:Immunoblotting; D045342:Light-Harvesting Protein Complexes; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009249:NADP / Q000378:metabolism; D010788:Photosynthesis / Q000235:genetics*; D045322:Photosynthetic Reaction Center Complex Proteins / Q000032:analysis / Q000737:chemistry / Q000187:drug effects / Q000235:genetics* / Q000648:ultrastructure; D045331:Photosystem I Protein Complex; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012974:Sodium Iodide / Q000494:pharmacology
chemical_list: D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D045342:Light-Harvesting Protein Complexes; D045322:Photosynthetic Reaction Center Complex Proteins; D045331:Photosystem I Protein Complex; C078702:photosystem I, PsaI subunit; C082500:photosystem I, psaL subunit; D009249:NADP; D002918:Chymotrypsin; D012974:Sodium Iodide