Functionally important domains of the large hydrophilic loop of CP47 as probed by oligonucleotide-directed mutagenesis in Synechocystis sp. PCC 6803

Abstract

The chlorophyll a-binding protein CP47 serves as core antenna to photosystem II (PS II). The predicted topology of CP47 exhibits six membrane-spanning regions and a large hydrophilic loop (loop E) which roughly includes 200 residues (255-455) and is presumably exposed to the lumenal side of the thylakoid membrane. Several lines of experimental evidence suggest that loop E might be involved in binding or stabilizing functional manganese in the catalytic site of water oxidation or in interacting with the extrinsic PS II-O protein (the 33-kDa manganese-stabilizing protein). To scan loop E for functionally important domains, oligonucleotide-directed mutagenesis has been used to introduce deletions of 3-8 residues in conserved and charged regions of loop E. In addition, one single-site mutation of the only histidine present in loop E was created (H343L). Domains deleted in delta 1 (I265-F268), delta 2 (T271-K277), delta 4 (T304-L309), delta 5 (F311-N317), and delta 12 (D440-P447) are required for stable assembly of functional PS II complexes. Deletion of domains delta 3 (K277-E283) and delta 11 (R422-E428) significantly reduces the level of assembled PS II and impairs photoautotrophic growth and oxygen evolution. Deletion of domain delta 8 (A373-D380) enhances the susceptibility to photoinhibition. In contrast, deletion of domains delta 6 (G333-I336), delta 7 (K347-R352), delta 9 (V392-Q394), and delta 10 (D416-F420) and mutation of H343 to leucine do not seem to severly interrupt PS II structure and function, although all mutants exhibit a slightly decreased stability of PS II as compared to the wild type. Thus, selected domains of the large hydrophilic loop of CP47 are important for PS II structure and function. With respect to possible sites of interaction between loop E of CP47 and the extrinsic PS II-O protein, our results indicate that none of the deletions in the region from residue 330 to 420 (delta 6, delta 7, delta 8, delta 9, delta 10) completely interrupts a functional association of the manganese-stabilizing protein to PS II, although the binding characteristics might be changed in some cases.

Accumulation of the D2 Protein Is a Key Regulatory Step for Assembly of the Photosystem II Reaction Center Complex in Synechocystis PCC 6803

“…In the absence of CP47, D1 and D2 were expressed without protein accumulation, and CP43 associated with PSII in the later stage of the assembly process. Analysis of Synechocystis mutants lacking PSII subunits (23)(24)(25)(26)(27) largely confirmed results from Chlamydomonas. In the absence of CP43, subunits CP47, D1, and D2 accumulated.…”

Photosystem II Assembly in CP47 Mutant of Synechocystis sp. PCC 6803 Is Dependent on the Level of Chlorophyll Precursors Regulated by Ferrochelatase

“…28). Deletion of four amino acid residues in the ⌬1 domain resulted in a non-photoautotrophic phenotype (17) and the complete loss of CP47 protein from thylakoid membranes (Fig. 3).…”

Changes in the cyanobacterial photosynthetic apparatus during acclimation to macronutrient deprivation

“…The peak at 650 nm is emitted from phycocyanin in the PBS. The peak at 685 nm is a combination of emissions from the terminal energy acceptors of the PBS (around 680 nm; Gantt and Lipschultz 1973;Gantt et al 1979) and from CP43, one of the Chl-binding proteins associated with PS II (around 686 nm; Pakrasi et al 1985;Dzelzkalns and Bogorad 1989;Nilsson et al 1992;Haag et al 1993 . 1992).…”