Isolation and Purification of Lipase from the Riboflavin Overproducing Fungus Eremothecium Ashbyii

Abstract

An extracellular lipase was observed in the hemiascomycete riboflavin overproducing fungus Eremotheciumashbyii . It was isolated and purified by salting out, dialysis, ion exchange chromatography on DEAE Sephadex and FPLC to 75-fold purity. The molecular weight of the purified enzyme was estimated to be 66 KDa and it showed a preference for unsaturated fatty acids. Maximum lipase was induced on lipid substrates containing a large percentage of unsaturated fatty acids. Thus olive oil substrate induced maximum lipase activity. It had an optimum pH of 7.0, an optimum temperature of 30°C, was stimulated by Zn 2+ , Mg 2+ , Fe 3+ , Ca 2+ and inhibited byCu 2+ , Co 2+ . An inhibition of activity upon incubation with PMSF and guanidine chloride, suggested the involvement of Serine and Arginine respectively in catalysis, whereas absence of inhibition by mercapto ethanol ruled out involvement of thiol groups in catalysis. This is the first report on the isolation and characterization of the lipase from the riboflavin over producer E. ashbyii. The possibility of lipase providing precursors for riboflavin over production can be exploited further to improve riboflavin production.