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Title:Experimental and theoretical studies of the photosystem II reaction center: Implications for bicarbonate binding and function
Author(s):Xiong, Jin
Doctoral Committee Chair(s):Govindjee
Department / Program:Biology
Degree Granting Institution:University of Illinois at Urbana-Champaign
Subject(s):Biology, Botany
Biophysics, General
Biology, Plant Physiology
Abstract:To investigate the role of arginine residues in bicarbonate binding in the photosystem II (PSII) reaction center, arginines 257 and 269 of the D1 polypeptide (D1-R257 and D1-R269) near the putative bicarbonate binding sites, the non-heme iron and the Q$\sb{\rm B}$ binding niche, were mutated in a unicellular green alga Chlamydomonas reinhardtii. The effects of mutations on bicarbonate binding and on other aspects of PSII photochemistry were investigated.
D1-R269 was mutated into a glycine. The mutant was shown to be unable to grow photoautotrophically and to have significant modifications on both the donor and acceptor sides of PSII, which includes a substantial reduction of the electron transfer kinetics from Q$\sb{\rm A}\sp{-}$ to the plastoquinone pool, a significantly increased minimal fluorescence level (F$\sb{\rm o})$ interpreted by a decrease in the excitation energy transfer from antennae to the PSII reaction center, a loss of the two-electron gate functioning, a reduction in 77 K PSII chlorophyll a fluorescence emission bands (F685 and F695) suggesting a possible destabilization of the PSII complex, a significantly modified Q$\sb{\rm B}$ niche, substantially reduced Tyr$\sb{\rm D}\sp+$/Q$\sb{\rm A}\sp-$ and Q$\sb{\rm A}\sp-$ Fe$\sp{2+}$ EPR signals, the absence of the S$\sb2$Q$\sb{\rm A}\sp-$ and S$\sb2$Q$\sb{\rm B}\sp-$ thermoluminescence bands in the mutant, the absence of the tetranuclear Mn cluster, and a 4 fold less sensitivity to the bicarbonate-reversible formate inhibition for the electron transfer from Q$\sb{\rm A}\sp-$ to the plastoquinone pool. However, the formate/bicarbonate binding site still appears to exit, suggesting that D1-R269 plays some role in the binding niche of bicarbonate but is of more importance for the general structure and function of PSII.
D1-R257 was mutated into a glutamate and a methionine. The mutants are able to grow photosynthetically, but at a slower rate compared to the wild type. Inhibition of the PSII reaction in the two mutants appears to be mainly on the acceptor side. The mutants show a significantly elevated F$\sb{\rm o}$ level, a reduction of 77 K fluorescence emission bands (F685 and F695), a reduction of PSII electron transfer, particularly, from Q$\sb{\rm A}\sp-$ to the plastoquinone pool. Although the binding affinity to the herbicide DCMU in the mutants appears to be unaltered, their sensitivity to the bicarbonate-reversible formate inhibition is drastically reduced. Further, the two mutants are poorly inhibited by arginine-specific reagents (phenylglyoxal and 2,3-butanedione) which inhibit the electron transfer in the wild type with a reversal by the addition of bicarbonate. The evidence implicates the near absence of bicarbonate binding in these two mutants and the essentiality of D1-R257 for the in vivo binding of bicarbonate/formate.
To obtain a more detailed understanding of the mechanism of bicarbonate-mediated PSII electron transfer as well as the structure of the PSII reaction center, three dimensional models of the PSII reaction center of C. reinhardtii and the cyanobacterium Synechocystis sp. PCC 6803 were generated. The models were constructed partly based on the significant homology with the anoxygenic purple bacterial photosynthetic reaction centers for which high resolution X-ray crystal structures are available, and partly based on existing experimental evidence. (Abstract shortened by UMI.)
Issue Date:1996
Rights Information:Copyright 1996 Xiong, Jin
Date Available in IDEALS:2011-05-07
Identifier in Online Catalog:AAI9712487
OCLC Identifier:(UMI)AAI9712487

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