The cysteine-proximal aspartates in the Fx-binding niche of photosystem I. Effect of alanine and lysine replacements on photoautotrophic growth, electron transfer rates, single-turnover flash efficiency, and EPR spectral properties.
The FX electron acceptor in Photosystem I (PS I) is a highly electronegative (Em = -705 mV) interpolypeptide [4Fe-4S] cluster ligated by cysteines 556 and 565 on PsaB and cysteines 574 and 583 on PsaA in Synechocystis sp. PCC 6803. An aspartic acid is adjacent to each of these cysteines on PsaB and adjacent to the proline-proximal cysteine on PsaA. We investigated the effect of D566PsaB and D557PsaB on electron transfer through FX by changing each aspartate to the neutral alanine or to the positively charged lysine either singly (D566APsaB, D557APsaB, D566KPsaB, and D557KPsaB) or in pairs (D557APsaB/D566APsaB and D557KPsaB/D566APsaB). All mutants except for D557KPsaB/D566APsaB grew photoautotrophically, but the growth of D557KPsaB and D557APsaB/D566APsaB was impaired under low light. The doubling time was increased, and the chlorophyll content per cell was lower in D557KPsaB and D557APsaB/D566APsaB relative to the wild type and the other mutants. Nevertheless, the rates of NADP+ photoreduction in PS I complexes from all mutants were no less than 75% of that of the wild type. The kinetics of back-reaction of the electron acceptors on a single-turnover flash showed efficient electron transfer to the terminal acceptors FA and FB in PS I complexes from all mutants. The EPR spectrum of FX was identical to that in the wild type in all but the single and double D566APsaB mutants, where the high-field resonance was shifted downfield. We conclude that the impaired growth of some of the mutants is related to a reduced accumulation of PS I rather than to photosynthetic efficiency. The chemical nature and the charge of the amino acids adjacent to the cysteine ligands on PsaB do not appear to be significant factors in the efficiency of electron transfer through FX.
Vassiliev
I R
IR
Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA.
Yu
J
J
Jung
Y S
YS
Schulz
R
R
Ganago
A O
AO
McIntosh
L
L
Golbeck
J H
JH
D000409:Alanine / Q000378:metabolism*; D019943:Amino Acid Substitution; D001224:Aspartic Acid / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites; D000458:Cyanobacteria / Q000378:metabolism*; D003545:Cysteine / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D007700:Kinetics; D008239:Lysine / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009249:NADP / Q000378:metabolism; D045322:Photosynthetic Reaction Center Complex Proteins / Q000378:metabolism*; D045331:Photosystem I Protein Complex; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D008565:Membrane Proteins; C073973:PSI-A protein, Synechococcus; D045322:Photosynthetic Reaction Center Complex Proteins; D045331:Photosystem I Protein Complex; C089909:photosystem I, psaB subunit; D001224:Aspartic Acid; D009249:NADP; D008239:Lysine; D003545:Cysteine; D000409:Alanine