title: Heterologous expression of lipases YLIP4, YLIP5, YLIP7, YLIP13, and YLIP15 from Yarrowia lipolytica MSR80 in Escherichia coli: Substrate specificity, kinetic comparison, and enantioselectivity.
abstract: Five lipase genes, ylip4, ylip5, ylip7, ylip13, and ylip15, from Yarrowia lipolytica MSR80 were cloned and expressed in the pEZZ18-HB101 system. The lipases shared maximum sequence identity with Candida galli lipase, whereas they shared structural similarity with YLIP2 of Y. lipolytica CLIB122. The enzymes, purified using IgG sepharose, had specific activities in the range of 7-25 U mg-1. Biochemical characteristics of all the lipases varied with respect to thermostability, substrate specificity, and enantioselectivity. All the enzymes were most active at neutral or slightly alkaline pH and were stable in the pH range 3.0-8.0, except YLIP4, which showed 50% stability at pH 10.0. Temperature optima of all the lipases varied from 30 to 50 ºC. YLIP15 and YLIP13 were most thermostable with a t1/2of 138 and 112 Min, respectively, at 60 °C. The lipases exhibited varied substrate specificity on p-nitrophenyl esters ranging from short-chain specificity (YLIP15), mid-chain specificity (YLIP4, YLIP5, YLIP7), and long-chain specificity (YLIP13). Catalytic efficiency on p-nitrophenylcaprate was highest for YLIP13 (67 × 103mM-1min-1) and lowest for YLIP15 (6.7 × 103mM-1min-1). YLIP13 was S-enantioselective, and YLIP15 was R-enantioselective with enantiomeric excess of 53 and 36%, respectively. Of all five lipases, YLIP13 and YLIP15 could be considered as industrially important enzymes as they were thermostable and enantioselective.
authors: Department of Microbiology, University of Delhi South Campus, New Delhi, India.
authors: Poonam
authors: P
authors: Syal
authors: Department of Microbiology, University of Delhi South Campus, New Delhi, India.
authors: Rani
authors: R
authors: Gupta
mesh_terms: D055162:Biocatalysis; D003001:Cloning, Molecular; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008049:Lipase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D019216:Metals, Heavy / Q000494:pharmacology; D008958:Models, Molecular; D055550:Protein Stability; D013237:Stereoisomerism; D013379:Substrate Specificity; D013696:Temperature; D025062:Yarrowia / Q000201:enzymology*
chemical_list: D004791:Enzyme Inhibitors; D019216:Metals, Heavy; D008049:Lipase
keywords: Yarrowia lipolytica; catalytic efficiency; enantioselectivity; kinetics; thermostability