CRYSTAL-STRUCTURE OF 3-ISOPROPYLMALATE DEHYDROGENASE FROM THE MODERATE FACULTATIVE THERMOPHILE, BACILLUS-COAGULANS - 2 STRATEGIES FOR THERMOSTABILIZATION OF PROTEIN STRUCTURES

The crystal structure of 3-isopropylmalate dehydrogenase from the mode rate facultative thermophile Bacillus coagulans (BcIPMDH) has been det ermined by the X-ray method, BcIPMDH is a dimeric enzyme composed of t wo identical subunits, each of which takes an open alpha/beta structur e with 11 alpha-helices and 14 beta-strands. The polypeptide is folded into two domains. The first domain is composed of residues 1-101 and 257-356, and the second domain, of residues 102-256, The latter domain s of the two subunits are associated with one another by a dyad axis t o make the dimer, locally forming a beta-sheet and a four-helix bundle . As compared with the structure of the enzyme from the extreme thermo phile Thermus thermophilus (TtIPMDH), a new short beta-sheet (residues 329-330 and 340-341) absent in TtIPMDH is formed by the insertion of 5 residues in BcIPMDH. In terms of determinants for thermostabilizatio n, both consistent and inconsistent changes were found between the two enzymes. The regions including inconsistent changes are formed by dif ferent usages of the determinants for stabilizing the loops at differe nt levels, Those in BcIPMDH contain some structural redundancies in le ngth of amino acid sequence and flexibility of residues, which seem to be unnecessary for the enzymatic reaction, Such redundancies are also found in the primary structure of the enzyme of the mesophile Bacillu s subtilis, but these parts are more stabilized in BcIPMDH by hydrogen bonds and salt bridges, On the other hand, TtIPMDH is stabilized by r educing such redundant parts, This contrast suggests that different st rategies may be preferred for thermostabilization, depending on temper ature.