Towards Structural Determination of the Water-splitting Enzyme

Abstract

A photosystem II preparation from the thermophilic cyanobacterium Synechococcus elongatus, which is es- pecially suitable for three-dimensional crystallization in a fully active form was developed. The efficient puri- fication method applied here yielded 10 mg of protein of a homogenous dimeric complex of about 500 kDa within 2 days. Detailed characterization of the preparation demonstrated a fully active electron transport chain from the manganese cluster to plastoquinone in the QB binding site. The oxygen-evolving activity, 5000–6000 mol of O2/(hmg of chlorophyll), was the highest so far reported and is maintained even at temperatures as high as 50 °C. The crystals obtained by the vapor diffu- sion method diffracted to a resolution of 4.3 Å. The space group was determined to be P212121 with four photosys- tem II dimers per unit cell. Analysis of the redissolved crystals revealed that activity, supramolecular organi- zation, and subunit composition were maintained dur- ing crystallization.