| Research Abstract |
Thermoactinomyces vulgaris R-47 produces two α-amylases TVAI (637 amino acid residues, MW=71,OOODa) and TVAII (585 amino acid residues, MW = 67,500Da), and they can hydrolyze not only starch but also cyclodextrins (CDs), which are scarcely hydrolyzed by other a-amylases. The X-ray structures of inactive mutant TVAIIs (Glu354->Ala or Asp325→Asn) complexed with β-CD (E354A/ β-CD) methyl-β-CD (D325N/methyl-β-CD) and maltohexose (D325N/G6), have been determined, showing that Phe286 is an important residue for TVAII to recognize CDs, and that the binding mode ofG6 as a substrate is very similar to that observed in the other a-amylase and a-amylase family enzymes. Kinetic analyzes of the mutant TVAIIs, F286A, F286L, F286Y and F286W (Phe286→Ala, Leu, Tyr, Trp) have also been done, supporting results from X-ray structures. The X-ray structure of TVAI has been determined at 1.6 Å, showing that there are some differences in the structures between TVAI and TVAII, related to the substrate specificity and thermostability of the enzyme.
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