Molecular characterization and bioinformatics studies of a lipase from Bacillus thermoamylovorans BHK67

A bacterium isolated from a hot-water spring identified as Bacillus thermoamylovorans BHK67 successfully produced a thermotolerant extracellular alkaliphilic lipase. The lipase was purified to homogeneity by anion exchange chromatography with 15-fold purification and 12.1% yield. The lipase appeared to be a hexameric protein as it possessed a single band of Mr 25 kDa in SDS PAGE and 150 kDa in Native PAGE. DLS analysis of purified Bacillus thermoamylovorans BHK67 lipase (BTL) also showed the molecular integrity, homogeneity and stability of the enzyme. The purified lipase showed maximum activity at pH 7.5 with a half-life of 10.5 h at 55 degrees C. Kinetic study of purified lipase by Lineweaver-Burk plot provided K-m (7.7 mM), V-max (90.9 U/mL/min), K-cat (227.3 s(-1)) and K-spec (29.4mM s(-1)) for substrate p-nitrophenylpalmitate. The purified lipase also showed astonishing stability following exposure to ethanol, n-propanol, iso-propanol, n-butanol and DMSO. Amino acid characterization of BTL by MALDI-TOF-MS showed considerable resemblance with lysophospholipase L1 related esterase of Lactobacillus ozensis DSM 23829. Experimental coupled molecular modeling postulated a structure-activity correlation of BTL as a probable contender in degradation of xenobiotic compounds, biocatalysis, biotransformation of compounds, synthesis of optically active compounds, foodstuff industry, anticancer therapeutics etc. (C) 2017 Elsevier B.V. All rights reserved.