Overexpression, purification and characterization of organic solvent stable lipase from Bacillus licheniformis RSP-09.
The lipase gene (543 bp) from Bacillus licheniformis RSP-09, a thermophilic isolate, was overexpressed in Escherichia coli BL21 (DE3). It encodes a polypeptide of 181 residues and has 96% identity with Bacillus pumilus B26 lipase gene. The recombinant lipase was purified 19-fold to electrophoretic homogeneity by His-tag chromatography. The molecular mass of the purified recombinant B. licheniformis RSP-09 lipase was found to be 24 kDa. The purified recombinant B. licheniformis RSP-09 lipase exhibited optimal activity at pH 10.0 and 40 degrees C. The apparent K(m) and V(max) values for pNPP were found to be 453 +/- 118 microM and 288.5 +/- 33.67 micromol min(-1) mg protein(-1), respectively. The purified recombinant lipase had a wide range of substrate specificity and exhibited tolerance to both detergents and organic solvents. Thus, enzyme has potential to be employed in detergents and biocatalysis in nonaqueous solvents.
Madan
Bhawna
B
Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology Delhi, Hauz Khas, New Delhi, India.
Mishra
Prashant
P
D001407:Bacillus / Q000201:enzymology*; D002846:Chromatography, Affinity / Q000379:methods; D003001:Cloning, Molecular; D003902:Detergents / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008049:Lipase / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012997:Solvents / Q000494:pharmacology; D013379:Substrate Specificity; D013696:Temperature
D003902:Detergents; D004791:Enzyme Inhibitors; D012997:Solvents; D008049:Lipase