THE ROLE OF PROTEIN PURIFICATION IN THE STRUCTURE DETERMINATION OF AN INTEGRAL MEMBRANE PROTEIN. G. McDermott⁺, S.M. Prince, M.Z. Papiz^*, A.M. Hawthornthwaite-Lawless^*, A.A. Freer, N.W. Isaacs, R.J. Cogdell^#. Dept.'s of Chemistry and ^#Biochemistry, University of Glasgow, Glasgow, G12 8QQ, UK. and; ^*CLRC Daresbury Laboratory, Daresbury, Warrington, WA4 4AD, UK. ⁺Present address: Dept. of Chemistry, University of Crete, Iraklion, Greece

Structural elucidation of the Nonameric LH2 complex, an integral membrane protein from the photosynthetic purple bacterium Rps. acidophila, required more than ten years work. For a large proportion of this time progress was frustrated by the poor "quality" of diffraction exhibited by crystals of the complex. In the initial stages of the analysis this was manifest in low resolution and poor reproducibility of diffraction. Latterly, when this problem had been alleviated, it became apparent that the level of isomorphism between "native" crystals was low. Clearly, a deleterious factor which rendered the search for heavy atom derivatives somewhat ambiguous.

Optimisation of diffraction, both in terms of maximum observed resolution and degree of isomorphism between "native" crystals, was a dynamic and ongoing process. This presentation will describe the evolution of the purification and crystallisation protocols and relate protocol changes to enhancement of diffraction quality.

The essence of this presentation will be derived from the distillation of a large volume of empirical observation. Consequently, some tentative proposals on diffraction improvement stratagems, potentially applicable to other membrane protein systems, will also be presented