MOLECULAR REPLACEMENT STUDIES OF A NUCLEASE INHIBITOR PROTEIN. C. Dennis`, R. Pauptit*, A. Tucker*, R.James`, C. Kleanthous`, G. Moore` and M. Osbourne`. `School of Biological Sciences, `University of East Anglia, Norwich, UK; *Zeneca Pharmaceuticals, Alderley Park, Macclesfield,Cheshire, UK

Structural studies are being carried out on a group of homologous nuclease inhibitor proteins using an available NMR structure as a trail modelin molecular replacement. These inhibitor proteins bind tightly to the nuclease and neutralise their toxicity. The colicin E family are a group of bacteriocins which are secreted as part of a bacterial defence mechanism which are induced by DNA damaging agents, such as UV light and are secreted as part of a bacterial defence mechanism. Colicin E2, E7, E8 and E9 exert toxicity by a Dnase activity in their C terminal domain. This toxicity is neutralised inside the producing cell by the tight binding of a constitutively expressed 10kDa immunity protein. This immunity protein shows high specificity to its cognate colicin. The NMR structure of Immunity protein 9 (Im9) is mainly helical with large flexible loops. Crystallisation trials have been successful for Im7 and large orthorhombic crystals diffracting to 2.0Å have been obtained. The immunity proteins share 60% homology so molecular replacement studies of Im7 are currently underway using the NMR structure of Im9 as the trial model.