Purification and characterization of a hyperthermostable and high maltogenic alpha-amylase of an extreme thermophile Geobacillus thermoleovorans.
The purified alpha-amylase of Geobacillus thermoleovorans had a molecular mass of 26 kDa with a pI of 5.4, and it was optimally active at 100 degrees C and pH 8.0. The T 1/2 of alpha-amylase at 100 degrees C increased from 3.6 to 5.6 h in the presence of cholic acid. The activation energy and temperature quotient (Q 10) of the enzyme were 84.10 kJ/mol and 1.31, respectively. The activity of the enzyme was enhanced strongly by Co2+ and Fe2+; enhanced slightly by Ba2+, Mn2+, Ni2+, and Mg2+; inhibited strongly by Sn2+, Hg2+, and Pb2+, and inhibited slightly by EDTA, phenyl methyl sulfonyl fluoride, N-ethylmaleimide, and dithiothreitol. The enzyme activity was not affected by Ca2+ and ethylene glycol-bis (beta-amino ethyl ether)-N,N,N,N-tetra acetic acid. Among different additives and detergents, polyethylene glycol 8000 and Tween 20, 40, and 80 stabilized the enzyme activity, whereas Triton X-100, glycerol, glycine, dextrin, and sodium dodecyl sulfate inhibited to a varied extent. alpha-Amylase exhibited activity on several starch substrates and their derivatives. The K m and K cat values (soluble starch) were 1.10 mg/ml and 5.9 x 10(3)/min, respectively. The enzyme hydrolyzed raw starch of pearl millet (Pennisetum typhoides) efficiently.
Uma Maheswar Rao
J L
JL
Department of Microbiology, University of Delhi South Campus, Benito Juarez Road, New Delhi, 110 021, India.
Satyanarayana
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D001406:Bacillaceae / Q000145:classification / Q000201:enzymology* / Q000648:ultrastructure; D004789:Enzyme Activation; D004795:Enzyme Stability; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000302:isolation & purification* / Q000648:ultrastructure; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008670:Metals / Q000378:metabolism; D008855:Microscopy, Electron, Scanning; D031782:Pennisetum / Q000378:metabolism; D013213:Starch / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
D008670:Metals; D013213:Starch; D006026:Glycoside Hydrolases; C120909:glucan 1,4-alpha-maltohydrolase