Open AccessOxygen evolution from water. Potential metal ligands in photosynthetic water oxidation identified by site‐directed mutagenesis of Synechocystis sp. PCC 6803
Author(s) -
Bowlby Neil R.,
Sithole Idah,
Babcock Gerald T.,
McIntosh Lee
Publication year - 1996
Publication title -
berichte der bunsengesellschaft für physikalische chemie
Language(s) - English
Resource type - Journals
ISSN - 0005-9021
DOI - 10.1002/bbpc.19961001209
Subject(s) - mutant , mutagenesis , chemistry , photosystem ii , thylakoid , site directed mutagenesis , manganese , biochemistry , synechocystis , photosynthesis , oxygen evolution , active site , biophysics , stereochemistry , biology , chloroplast , enzyme , gene , organic chemistry , electrode , electrochemistry
In order to identify amino acids that could serve as ligands to the manganese cluster, which comprises the active site of the oxygen‐evolving complex (OEC), and the calcium ion that affects the stability of the OEC, we have generated a series of site‐directed mutants in the D1 polypeptide of the PS II reaction center. Our approach has been to emphasize characterization of mutants with reduced PS II activity rather than those that lack PS II activity altogether in order to address questions about the effects of the mutations on the functional properties of the Mn ensemble. A number of mutants have been isolated with impaired PS II activity and a characterization of in vivo photosynthesis from mutant and WT strains is presented. All of the mutants described in this report produce nearly WT levels of PS II in the thylakoid membrane. Non‐conservative mutation at Asp‐103 (DN103D1), Glu‐104 (EQ104D1) or Glu‐333 (EQ333D1) results in moderate (∼ 50%) inhibition of oxygen evolution, whereas mutation at Glu‐65 (EQ65D1) or His‐337 (HF337D1) results in cells that retain only 10 to 20% of WT activity. In vivo fluorescence induction kinetics are consistent with a lesion on the donor side of PS II in each of the mutants tested. We conclude that Glu‐65 and His‐337 are directly involved in binding manganese or calcium, and that Asp‐103 and Glu‐104 may be involved in calcium binding but are not manganese ligands.