Fervidobacterium changbaicum Lip1: identification, cloning, and characterization of the thermophilic lipase as a new member of bacterial lipase family V

• DOI: 10.1007/s00253-010-2971-y
• Publication Date: 2010-11-02T15:32:38Z
• Authors: Jingang Cai; Yuan Xie; Bo Song; Yanping Wang; Zuoming Zhang; Yan Feng
• Keywords: 0301 basic medicine; Hot Temperature; Bacteria; Sequence Homology, Amino Acid; Molecular Sequence Data; Lipase; Sequence Analysis, DNA; Hydrogen-Ion Concentration; Recombinant Proteins; Substrate Specificity; Kinetics; 03 medical and health sciences; Enzyme Stability; Amino Acid Sequence; Cloning, Molecular; Phylogeny

A novel lipase gene encoded 315 amino acid residues was obtained using lipase-prospecting primers and genome walking from hyperthermophilic bacterium Fervidobacterium changbaicum CBS-1. Sequence alignment and phylogenetic analysis revealed this novel lipase is a new member of bacterial lipase family V. The recombinant enzyme F. changbaicum lipase 1 (FCLip1) showed maximum activity at 78 °C and pH 7.8. It displayed extreme thermostability at 70 °C and was also stable across a wide pH range from 6.0 to 12.0. Kinetic study demonstrated FCLip1 preferentially hydrolyzed middle-length acyl chains, especially p-nitrophenyl caprate and tricaprylin. With p-nitrophenyl caprate as a substrate, the enzyme exhibited a K(m) and k(cat) of 4.67 μM and 22.7/s, respectively. In addition, FCLip1 was resistant to various detergents and organic solvents. This enzyme is the first reported thermophilic lipase from bacterial family Thermotogaceae. Its extreme stability with respect to temperature and pH, along with its triglyceride hydrolysis activity, indicate that FCLip1 has high potential for future application.