STRUCTURE SOLUTION OF AN INTEGRAL MEMBRANE PROTEIN: NOVEL DERIVITIZATION METHODS. S.M. Prince, M.Z. Papiz^*, G. Mcdermott, A.M. Hawthornthwaite-Lawless^*, A.A. Freer, R.J. Cogdell^~, N.W. Isaacs. Dept.'s of Chemistry and ^~Biochemistry, University of Glasgow, Glasgow, G12 8QQ, UK. and; ^*CLRC Daresbury Laboratory, Daresbury, Warrington, WA4 4AD, UK.

The structure of the LH2 complex from purple bacteria was pursued using a number of novel derivitization techniques. The crystal structure of this integral membrane complex revealed a highly symmetrical molecule of nine subunits possessing C9 molecular symmetry, the C9 axis being coincident with the 3-fold axis of the R32 spacegroup. A number of factors obstructed or confused standard MIR techniques. These include; the small extent of the polar surface of the molecule, the alignment of the molecule with the crystallographic 3-fold - resulting in multiple sites with the same z fractional coordinate and the high pH (9.3) at which crystals were stable. These compromised; derivative binding, Patterson solution, and heavy atom salt solubility.

A number of techniques were attempted to form isomorphous heavy atom derivatives including: Seleno-methionine labelling and the exchange of bacteriochlorophyll co-factors for Pd substituted analogues. The method which finally resulted in a phase set used traditional heavy atom salt soaks with a additional stage. The second stage simply employed differing binding site dynamics or differing heavy atom salt chemistry to partially 'back-soak' away a subset of sites. This resulted in a considerable gain in isomorphism and difference Patterson maps trivial to solve.

The presentation will describe these derivitization methods, and successive manipulations demonstrating the quality of the phase set obtained.