Characterization of a heterotrophic mutant of Synechocystis sp. PCC 6803 created by random mutagenesis targeted to the psbAII gene

摘要

The photosystem II (PSII) is unique in that it can generate a high redox potential to oxidize water molecules. To elucidate molecular architectures behind the unique function, the structure of PSII complex has been analyzed extensively during the past two decades. However, the structural information provided by x-ray crystallographic analysis at this moment is insufficient to locate amino acid residues in the complex and to understand the molecular details (Zouni A. et al. 2001). The mutational analysis, on the other hand, has a potential to provide specific information on the function of particular amino acid residues and the structure of complex. Thus we have been engaged in random mutational analysis targeted to the D1 subunit, in order to discover unexpected structure-function relationships in the PSII reaction center, both in static and dynamic aspects. By this method, we have created more than one hundred random mutants impaired in the capacity of photoautotrophic growth caused by 1-9 aminoacid substitution(s) in the targeted 210 amino acid (Ser148-Ala357) region of the D1 protein (See, Yamasato et al., S22-029). This article provides a preliminary characterization of a mutant (a triple mutant named npRK07) obtained by this kind of randommutagenesis, which carries 3 amino acid substitutions on the cytosolic side of D1 protein, i.e., F255I, Q261P and H272R. We have generated 3 single-point mutants, each possessing one of three substitutions in the triple mutant, and demonstrated that theH272R substitution at the putative ligand for non-heme Fe on the acceptor side dramatically affects the structure and function on the donor side in the PSII complex and is responsible for the observed loss of photoautotrophy in the random mutant.