Isolation and Structural Characterization of Monomeric and Trimeric Photosystem I Complexes (P700·FA/FB and P700·Fx) from the Cyanobacterium Synechocystis PCC 6803

Abstract

An isolation procedure was developed for the cyanobacterium Synechocystis 6803 (and 6714) which yields both monomeric and trimeric photosystem I complexes (P700·FA/FB complexes) depleted of the stroma-exposed subunits PsaC, -D, and -E (P700·FX complexes). Analysis by high resolution gel electrophoresis in combination with immunoblotting and N-terminal sequencing reveals the selective and quantitative removal of PsaC, -D, and -E from the P700·FA/FB complex, containing PsaA, -B, -C, -D, -E, -F, -K, -L and at least two subunits ≤4 kDa. Monomeric and trimeric P700·Fx complexes show an identical subunit composition and an identical charge recombination half-time of 750 ± 250 µs as determined by flash-induced absorption change measurements, reflecting the quantitative loss of iron-sulfur clusters FA/FB and the presence of cluster FX. The existence of a stable trimeric P700·FX complex enables a detailed structural analysis by electron microscopy with high resolution. Comparison of averaged top and side view projections of P700·FX and P700·FA/FB complexes show that the height of the complex is reduced by about 2.5-3.3 nm upon removal of the three stroma-exposed subunits and indicate the position of these three subunits on the PS I surface. While the outer contours of the stroma exposed mass of PS I agree very well with the three-dimensional crystal analysis recently published for trimeric PS I of Synechococcus elongatus, only the structural analysis presented here is able to assign the stroma-exposed mass exclusively to the subunits PsaC, -D, and -E and to exclude a contribution of other subunits.