Open AccessPurification and Characterization of a Noble Thermostable Alpha-amylase from Anoxybacillus tengchongensis RA1-2-1 Isolated from Geothermal Spring of Nepal
Author(s) -
Parash Mani Timilsina,
Gyanu Raj Pandey,
Asmita Shrestha,
Manish Ojha,
Garima Baral,
Tika Bahadur Karki
Publication year - 2020
Publication title -
journal of food science and technology nepal
Language(s) - English
Resource type - Journals
ISSN - 1816-0727
DOI - 10.3126/jfstn.v12i12.31038
Subject(s) - thermophile , hot spring , strain (injury) , enzyme , 16s ribosomal rna , hydrolysis , amylase , chemistry , bacteria , enzyme assay , microbiology and biotechnology , chromatography , food science , biochemistry , biology , paleontology , genetics , anatomy , gene
A thermophilic amylolytic strain, Anoxybacillus tengchongensis RA1-2-1 was isolated from geothermal spring of Rasuwagadi district of Nepal. The BLAST alignment of the 16s rRNA sequence revealed 99.3% similarity with the type strain Anoxybacillus tengchongensis T-11. The morphological, physiological and biochemical properties were similar to the type strain. The enzyme from the strain was purified to 40-fold purification by DEAE-cellulose ion exchange chromatography. The Km value of the enzyme was 0.68±0.05 mg/ml. The optimum pH and temperature were 7.0 and 70 °C. SDS-PAGE analysis showed a single band at 69 kDa. The half-life of the enzyme at 70°C and 80°C were 85.01min and 51.96 min respectively. TLC analysis of the hydrolysis product showed that the enzyme is maltogenic amylase. The calcium independent enzyme was completely inhibited by Hg2+ but showed inhibitory effect in the range of 100 %-30 % in the presence of other salts at 1-10 mM concentrations.