The PsbW-protien; Its location and involvement in photoinhibition

Abstract

Roughly half of the photosystem II polypeptides are encoded by the nucleus while the reaction centre core components are chloroplast encoded. The nuclear encoded photosynthetic proteins are synthesised in the cytosol with N-terminal extensions that direct the proteins to the right compartment in the cell. Various biochemical and crystallographic studies of PSII have shown that the assembled PSII-complex in vivo is a dimer. Building a functional PSII thus not only involves targeting to the envelope and the thylakoid membrane as well as processing and assembly of monomers, but also dimerization and lateral migration. Recent data strongly suggest that the nuclear encoded, 6.1 kDa psbW-protein takes part in the dimerization. We have combined an experimental system in which nuclear encoded radiolabelled precursor proteins are incubated with isolated chloroplasts, with blue native PAGE. The blue native tris/tricine system resolves nine distinct bands of which three correspond to PSII dimers/supercomplexes. Using this combined assay, kinetic import/assembly studies show that all so far analysed thylakoid membrane proteins are found initially in the stroma lamellae fractions before migration to their proper locations. It was found that the time for the PsbW-protein, from envelope targeting to incorporation into a fully assembled dimeric PSII, is less than three minutes. This In Organello assay in combination with site directed mutagenesis has also proven to work as a tool to analyse the importance of specific amino acid residues with respect to the PSII dimerization. The PsbW-protein N-terminal domain is being analysed and will be discussed.