PARENT SESSION
Posters P2A Type I reaction centres. Abstracts (181-218)

The role of the asymmetric protein environment upon the properties of P₇₀₀. Kevin Redding^*,1, Maria Pantelidou², Tatyana Konovalova¹, Johan van Tol³, Fabrice Rappaport⁴, Gary Hastings⁵, Brian Abbott¹, Alexander Petrenko¹, ¹ The University of Alabama, Tuscaloosa, Alabama, USA² Iowa State University, Ames, Iowa, USA³ National High Magnetic Field Laboratory, Tallahassee, Florida, USA⁴ Institut de Biologie Physico-Chimique, Paris, France⁵ Georgia State University, Atlanta, GA, USA

ABSTRACT- P₇₀₀, the presumed primary electron donor of Photosystem I (PS1), is a heterodimer of chlorophyll a (P_B) and a′ (P_A). The PS1 crystal structure reveals that P_A is H-bonded to a Thr residue, which is part of a larger H-bonding network consisting of a Ser, Tyr and tightly-bound water molecule. There is no such network near P_B, where the corresponding residues are a Tyr, Gly, and Leu, respectively. To investigate the influence of the H-bond to P_A, PsaA-Thr739 was converted to Ala in Chlamydomonas reinhardtii. The mutant PS1 still contained 1 Chl a′ per PS1, but the mutation provoked significant differences in the P₇₀₀/P₇₀₀⁺ visible and FTIR difference spectra and a 60-mV decrease in the P₇₀₀/P₇₀₀⁺ midpoint potential. FTIR and ¹H-ENDOR analysis were consistent with a slightly altered charge and unpaired spin distribution over the chlorophylls in P₇₀₀⁺, with 8-16% of the charge/spin on P_B relocated onto P_A. The mutation did not cause an observable change in the directionality of electron transfer within PS1. More comprehensive multiple mutants have now been constructed in Synechocystis PCC6803, in which the three main residues of the H-bonding network near P_A have been converted to the corresponding residues on the P_B side and vice versa. Interestingly, while the A-side triple mutation had the most profound effect upon the visible difference spectrum, the B-side triple mutation produced the largest shift in redox potential (35-mV increase). The A-side triple mutant displayed an increase in the hyperfine coupling of the 12-methyl protons, and this change was tied to the Thr mutation, since a PsaA-T743F mutant produced an indistinguishable ¹H-ENDOR spectrum. In both cases, there was still 1 Chl a′ per PS1. Surprisingly, the B-side mutations (PsaB-Y727T, PsaB-L590Y/G594S, and triple mutant) displayed unexpectedly large shifts in hyperfine couplings and changes in the g-tensor of P₇₀₀⁺.

KEY WORDS: site-directed mutagenesis, photosystem I, asymmetry