α‐Amylases from Thermoactinomyces vulgaris: characteristics, primary structure and structure prediction

Abstract

Two amylolytic active protein fractions (named α‐amylase 1 and α‐amylase 2) were isolated from the bacterium Thermoactinomyces vulgaris strain 94‐2A. α‐Amylase 1 had a molecular mass of 51.6 kDa, whereas α‐amylase 2 consists of two fragments which have molecular masses of 17.0 and 34.6 kDa, respectively. These two fragments are products from a proteolytic cleavage of a‐amylase 1 at amino acid position 303 (tryptophan) by a serine protease (thermitase) which is also produced by T. vulgaris. The purified α‐amylase 1 and 2 follow the Michaelis‐Menten kinetics in the presence of starch as substrate with Km values of 1.37 ± 0.07 and 1.29 ± 0.18 mg/mL, respectively. In effect they differ in their stability characteristics. The amino acid sequence of α‐amylase from T. vulgaris derived from DNA sequence (1) was compared with those of other α‐amylases. It reveals high homologies to α‐amylases from other microorganisms (e.g. B. polymyxa, A. oryzae, S. occidentalis and S.fibuligera). A three‐dimensional structure model for α‐amylase 1 on the basis of the 3 Å X‐ray structure of Taka‐amylase was constructed.

Gene sequence, modeling, and enzymatic characterization of α-amylase AmyLa from the thermophileLaceyellasp. DS3

“…The enzyme's V max did not change in the presence of glucose but K m increased to 1 mg/mL which confirms its competitive inhibition effect. This represents advantage to AmyLa α‐amylase, since the K m of most microbial α‐amylases including AmyTV1 is higher than 1 mg/mL.…”