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High-level Expression of Pro-Form Lipase From Rhizopus Oryzae in Pichia Pastoris and Its Purification and Characterization

High-level Expression of Pro-Form Lipase From Rhizopus Oryzae in Pichia Pastoris and Its Purification and Characterization

Jian-Rong Wang, Yang-Yuan Li, Shu-De Xu, Peng Li, Jing-Shan Liu, Dan-Ni Liu

Int J Mol Sci. 2013 Dec 24;15(1):203-17.

PMID: 24368519

Abstract:

A gene encoding Rhizopus oryzae lipase containing prosequence (ProROL) was cloned into the pPICZαA and electrotransformed into the Pichia pastoris X-33 strain. The lipase was functionally expressed and secreted in Pichia pastoris with a molecular weight of 35 kDa. The maximum lipase activity of recombinant lipase (rProROL) was 21,000 U/mL, which was obtained in a fed-batch cultivation after 168 h induction with methanol in a 50-L bioreactor. After fermentation, the supernatant was concentrated by ultrafiltration with a 10 kDa cut off membrane and purified with ion exchange chromatography using SP Sepharose Fast Flow chromatography. The optimum pH and temperature of the rProROL were pH 9.0 and 40 °C, respectively. The lipase was stable from pH 4.0 to 9.0 and from 25 to 55 °C. The enzyme activity was enhanced by Ca(2+) and inhibited by Hg(2+) and Ag(+). The lipase showed high activity toward triglyceride-Tripalmitin (C16:0) and triglyceride-Trilaurin (C12:0).

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP9001621-H	Lipase from Rhizopus oryzae		9001-62-1	Price

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