Purification and characterization of organic solvent stable protease from Bacillus licheniformis RSP-09-37.
A protease was purified from the cell-free supernatant of Bacillus licheniformis RSP-09-37, a mutant from a thermophilic bacterial strain, B. licheniformis RSP-09, using affinity chromatography with alpha-casein agarose resin. The protease was purified 85-fold to electrophoretic homogeneity. The apparent molecular mass of purified protease was 55 kDa using gel filtration in high-performance liquid chromatography, which is in agreement with the results obtained from sodium dodecyl sulfate-polyacrylamide gel electrophoresis, suggesting a monomeric nature of the protein. The purified protease revealed temperature optima of 50 degrees C and pH optima of 10.0 and was classified as serine protease based on its complete inhibition with phenyl methyl sulfonyl fluoride. The purified protease exhibited tolerance to both detergents and organic solvent. The synthetic activity of the protease was tested using the transesterification reaction between N-acetyl-L-phenylalanine-ethyl ester and n-propanol in organic solvents varying in their log P values and the kinetic parameters of the enzyme in these organic solvents were studied. The enzyme has potential to be employed for synthetic reactions and in detergent formulations.
Sareen
Ritu
R
Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology Delhi, Hauz Khas, New Delhi, India.
Mishra
Prashant
P
D001407:Bacillus / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D004795:Enzyme Stability; D004951:Esterification; D045482:Hot Springs / Q000382:microbiology; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009930:Organic Chemicals / Q000737:chemistry*; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D013696:Temperature
D001426:Bacterial Proteins; D009930:Organic Chemicals; D012697:Serine Endopeptidases