TWO STRATEGIES FOR THERMOSTABILIZATION IN 3-ISOPROPYLMALATE DEHYDROGENASE D. Tsuchiya, A. Takenaka, Department of Life Science, Tokyo Institute of Technology, Yokohama, Japan; T. Sekiguchi, Y. Nosoh, Department of Fundamental Science, Iwaki Meisei University, Fukushima, Japan.

Several factors have been proposed to account for the thermostability of proteins. For understanding the availabilities to thermal environments, it is necessary to reveal protein structures with intermediate properties between thermolabile and thermostable proteins. We have determined a crystal structure of 3-isopropylmalate dehydrogenase from the moderate facultative thermophile Bacillus coagulans (BcIPMDH) at 3.0 Å resolution with an R-factor of 0.185. A comparison of the structure with that of the enzyme from the extreme thermophile Thermus thermophilus (TtIPMDH) shows remarkable differences in the four loop regions though their overall tertiary structures are similar. These loops in BcIPMDH have structural redundancies in length of amino acid sequence, flexibility of residues and architecture of the tertiary structure, which seem to be unnecessary to retain the active site. Such redundancies are also found in the enzyme of the mesophile Bacillus subtilis, but those parts are more stabilized in BcIPMDH by hydrogen bonds and ion pairs. On the other hand, TtIPMDH is different in structural economy so that redundant parts are reduced to adapt at higher temperature. This contrast suggests the two different strategies depending on temperature. One is "reinforcement of structure", which has high enthalpic effects and results in minor changes of thermostabilization. The other is "reduction of structural redundancy", which is required when the former strategy is not sufficient to stabilize the structure at higher temperature.