Purification and characterization of two distinct lipases from Geotrichum candidum.
Lipase, an enzyme that hydrolyzes triacylglycerol, has been purified and characterized. The purification procedure includes ethanol precipitation and chromatographies on Sephacryl-200 HR, high resolution anion-exchange (mono Q) and Polybuffer exchanger 94. With this procedure, two forms of lipases from Geotrichum candidum were obtained. Lipase I (main enzyme) and lipase II (minor enzyme) were purified 35-fold with a 62% recovery in activity and 94-fold with a 18% recovery in activity, respectively. Their molecular weights have been estimated by polyacrylamide gel electrophoresis under denaturing conditions and by molecular sieving under native conditions at 56,000. Lipase I and II had optimum pH values of 6.0 and 6.8 and isoelectric points of 4.56 and 4.46, respectively. The enzymes are stable at a pH range of 6.0 to 8.0. Monovalent ions had little effect on both enzyme activities, while divalent ions at concentrations above 50 mM inhibited the lipase activities in a concentration-dependent manner. Sodium dodecyl sulfate at a concentration lower than 10 mM completely inhibited the lipase activity.
Veeraragavan
K
K
Protein Engineering Section, National Research Council of Canada, Montreal.
Colpitts
T
T
Gibbs
B F
BF
D000596:Amino Acids / Q000032:analysis; D002845:Chromatography; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D000431:Ethanol; D005848:Geotrichum / Q000201:enzymology*; D007525:Isoelectric Focusing; D008049:Lipase / Q000302:isolation & purification*; D003904:Mitosporic Fungi / Q000201:enzymology*; D008970:Molecular Weight
D000596:Amino Acids; D000431:Ethanol; D008049:Lipase