A thermostable lipolytic endospore forming Bacillus sp. strain L2 was isolated from a hot
spring (91oC) in Slim River, Perak, Malaysia. Nutritional studies showed that casamino
acid was the best nitrogen source while trehalose and Tween 60 were the best carbon
source and substrate respectively, for lipase production. The enzyme production was
promoted in the presence of metal ions such as Ca2+, Cu2+, Fe2+, Co2+, Zn2+, and Sr2+. The
optimum lipase production was obtained in the presence of CaCl2. Physiological study
indicate that, this strain showed optimal lipase production after 28 h of incubation at 70
oC with 1% starting inoculum. The enzyme was purified to homogeneity by ammonium
sulphate precipitation and affinity column chromatography (Heparin), with 3 fold
increase in specific activity and 48.9 % recovery. The purified enzyme produced a single
band on SDS-PAGE. Its molecular mass was estimated to be 43 KDa. The optimum pH
and temperature for L2 lipase were determined to be 7.0 and 80 oC, respectively. The
III
lipase was very stable for 2 h at 80 oC. The stability at higher temperature was shown to
be calcium dependent. The lipase activity was enhanced by Fe2+and Ca2+ while Cu2+
inhibited it. The enzyme activity was strongly inhibited by PMSF and completely
inhibited by pCMB. Substrates such as olive oil and sesame oil enhanced the lipase
activity
