PBR: a heavy-atom refinement and phasing procedure to reduce phase bias when heavy-atom derivatives contain common sites

A procedure, called PER (phase-bias reduction), has been developed to prope rly refine heavy-atom derivatives and to generate less biased heavy-atom ph ases when these derivatives contain common heavy-atom sites, Two independen t events are obtained by splitting the refinement and phasing calculations into two stages, the first in which one of the derivatives having common si tes is used together with the native amplitudes and the second in which bot h derivatives with common sites are used simultaneously, with one of them b eing used as the native data set. Improved centroid phases and the correspo nding figures of merit are obtained by phase combination. This procedure ha s been used in the structure determination of the iron-cluster-containing p rotein pyruvate-ferredoxin oxidoreductase. When the common heavy-atom sites are properly treated by the PER procedure, the resulting calculated centro id phases are improved with respect to classical heavy-atom refinement cent roid phases where all derivatives are refined together. This leads to impro ved electron-density distributions, since anomalous difference Fourier maps calculated with the PER-refined centroid phases and corresponding figures of merit show more clearly the positions of the iron sites.