Expression of Thermostable Lipase Gene from Bacillus stearothermophilus TP811 in E. coli M15 (pREP4)
Padchanee Sangthong*[a], Supachok Sinchaikul [b], Hataichanoke Niamsup [a], Shui-Tein Chen [b] and Suree Phutrakul [a]* Author for corresponding; e-mail address: padchanee1@hotmail.com
Volume: Vol.29 No.3 (DECEMBER 2002)
Research Article
DOI:
Received: 10 May 2002, Revised: -, Accepted: 9 October 2002, Published: -
Citation: Sangthong P., Sinchaikul S., Niamsup H., Chen S. and Phutrakul S., Expression of Thermostable Lipase Gene from Bacillus stearothermophilus TP811 in E. coli M15 (pREP4), Chiang Mai Journal of Science, 2002; 29(3): 161-171.
Abstract
The gene coding for a thermostable lipase of Bacillus stearothermophilus TP811 which was isolated from Teppanom hot spring in Chiang Mai was cloned into pQE 60 expression vector and transformed into E.coli M15(pREP4). Sequence analysis showed an open reading frame of 1254 bp encoding a polypeptide of 417 amino acid residues. Its extracellular lipase activity was higher than native strain about 43 times and about 2.6 times higher than pUC TP811 lipase (same lipase in pUC 19 vector). The lipase was most active at 45-55°C in alkaline condition of around pH 7.5-9, using p- nitrophenyl laurate (p-NPL) as synthetic substrate. The addition of 0.6 mM IPTG could induce the production of extracellular lipase about 2 times compared to non-induced condition. The addition of 1.0 g/l Tween 80 or 1.75 g/l Triton X-100 in LB medium could further increase the extracellular lipase production by 1.7 and 1.6 folds compared to 0.6 mM IPTG addition alone.