ENZYME ACTIVITY AT 100deg.C: CITRATE SYNTHASE FROM PYROCOCCUS FURIOSUS Rupert JM Russell, David W Hough, Michael J Danson and Garry L Taylor.School of Biology and Biochemistry,University of Bath, BA2 7AY, U.K.

The ability of enzymes from hyperthermophiles (growth temperature > 85deg.C) to be stable and optimally active at high temperatures has widespread biotechnological applications. In order to gain insights into enzyme mechanism at 100deg.C, the crystal structure of the closed form (liganded) of citrate synthase from the hyperthermophilic Archaeon Pyrococcus furiosus has been determined to 3.0.Å This has allowed direct structural comparisons between the same enzyme but from hosts which show different temperature optimum for citrate synthase activity: 37deg.C (pig), 55deg.C (Thermoplasma acidophilum) and now 100deg.C (P. furiosus). Pig citrate synthase undergoes a large conformational change upon substrate binding, but the structures of the thermophilic citrate synthases suggest that the magnitude of the conformational change may not be a constant feature of this enzyme mechanism. Coenzyme A is recognised in a slightly different manner from that observed in the pig enzyme. The differences in the active site and magnitude of the conformational change between the enzymes will be rationalised on the basis of performing the same catalytic mechanism but at different temperatures.