title: Cloning and characterization of two new thermostable and alkalitolerant α-amylases from the Anoxybacillus species that produce high levels of maltose.
abstract: Two genes that encode α-amylases from two Anoxybacillus species were cloned and expressed in Escherichia coli. The genes are 1,518 bp long and encode 506 amino acids. Both sequences are 98% similar but are distinct from other well-known α-amylases. Both of the recombinant enzymes, ASKA and ADTA, were purified using an α-CD-Sepharose column. They exhibited an optimum activity at 60°C and pH 8. Both amylases were stable at pH 6-10. At 60°C in the absence of Ca²⁺, negligible reduction in activity for up to 48 h was observed. The activity half-life at 65°C was 48 and 3 h for ASKA and ADTA, respectively. In the presence of Ca²⁺ ions, both amylases were highly stable for at least 48 h and had less than a 10% decrease in activity at 70°C. Both enzymes exhibited similar end-product profiles, and the predominant yield was maltose (69%) from starch hydrolysis. To the best of our knowledge, most α-amylases that produce high levels of maltose are active at an acidic to neutral pH. This is the first report of two thermostable, alkalitolerant recombinant α-amylases from Anoxybacillus that produce high levels of maltose and have an atypical protein sequence compared with known α-amylases.
authors.affiliation: Faculty of Biosciences and Bioengineering, Universiti Teknologi Malaysia, 81310 Skudai, Johor, Malaysia.
authors.forename: Yen Yen
authors.initials: YY
authors.lastname: Chai
authors.forename: Raja Noor Zaliha Raja Abd
authors.initials: RN
authors.lastname: Rahman
authors.forename: Rosli Md
authors.initials: RM
authors.lastname: Illias
authors.forename: Kian Mau
authors.initials: KM
authors.lastname: Goh
mesh_terms: D000595:Amino Acid Sequence; D058677:Anoxybacillus / Q000201:enzymology*; D001483:Base Sequence; D003001:Cloning, Molecular; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008320:Maltose / Q000378:metabolism*; D013213:Starch / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D000516:alpha-Amylases / Q000235:genetics / Q000378:metabolism*
chemical_list: D008320:Maltose; D013213:Starch; D000516:alpha-Amylases