Purification, crystallization and X-ray diffraction analyses of the T. elongatus PSII core dimer with strontium replacing calcium in the oxygen-evolving complex

Abstract

The core complex of photosystem II (PSII) was purified from thermophilic cyanobacterium Thermosynechococcus elongatus grown in Sr(2+)-containing and Ca(2+)-free medium. Functional in vivo incorporation of Sr(2+) into the oxygen-evolving complex (OEC) was confirmed by EPR analysis of the isolated and highly purified SrPSII complex in agreement with the previous study of Boussac et al. [J. Biol. Chem. 279 (2004) 22809-22819]. Three-dimensional crystals of SrPSII complex were obtained which diffracted to 3.9 A and belonged to the orthorhombic space group P2(1)2(1)2(1) with unit cell dimensions of a=133.6 A, b=236.6 A, c=307.8 A. Anomalous diffraction data collected at the Sr K-X-ray absorption edge identified a novel Sr(2+)-binding site which, within the resolution of these data (6.5 A), is consistent with the positioning of Ca(2+) in the recent crystallographic models of PSII [Ferreira et al. Science 303 (2004) 1831-1838, Loll et al. Nature 438 (2005) 1040-1044]. Fluorescence measurements on SrPSII crystals confirmed that crystallized SrPSII was active in transferring electrons from the OEC to the acceptor site of the reaction centre. However, SrPSII showed altered functional properties of its modified OEC in comparison with that of the CaPSII counterpart: slowdown of the Q(A)-to-Q(B) electron transfer and stabilized S(2)Q(A)(-) charge recombination.