Purification and characterization of an extracellular alpha-amylase from Bacillus subtilis AX20.
A Bacillus subtilis AX20 from soil with ability to produce extracellular alpha-amylases was isolated. The characterization of microorganism was performed by biochemical tests as well as 16S rDNA sequencing. Maximum amylase activity (38 U/ml) was obtained at stationery phase when the culture was grown at 37 degrees C. The enzyme was purified to homogeneity with an overall recovery of 24.2% and specific activity of 4133 U/mg. The native protein showed a molecular mass of 149 kDa composed of a homodimer of 78 kDa polypeptide by SDS-PAGE. The optimum pH and temperature of the amylase were 6 and 55 degrees C, respectively. The enzyme was inhibited by Hg(2+), Ag(2+), and Cu(2+) and it did not show an obligate requirement of metal ions. The enzyme was not inhibited by EDTA or EGTA, suggesting that this enzyme is not a metalloenzyme. The end products of corn starch and soluble starch were glucose (70-75%) and maltose (20-25%). Rapid reduction of blue value and the end products suggest an endo mode of action for the amylase. The purified amylase shows interesting properties useful for industrial applications.
Najafi
Mohsen Fathi
MF
Razi Vaccine and Serum Institute, Ahmadabad Street, Mashad, Iran. najafi99@yahoo.com
Deobagkar
Dileep
D
Deobagkar
Deepti
D
D001412:Bacillus subtilis / Q000201:enzymology*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification*; D004795:Enzyme Stability; D005947:Glucose / Q000096:biosynthesis; D006863:Hydrogen-Ion Concentration; D008320:Maltose / Q000096:biosynthesis; D019216:Metals, Heavy / Q000494:pharmacology; D013329:Structure-Activity Relationship; D013696:Temperature; D000516:alpha-Amylases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification*
D001426:Bacterial Proteins; D019216:Metals, Heavy; D008320:Maltose; D000516:alpha-Amylases; D005947:Glucose