[Cloning and expression of organic solvent tolerant lipase gene from Staphylococcus saprophyticus M36].
Lipases are important biocatalysts that are widely used in food processing and bio-diesel production. However, organic solvents could inactivate some lipases during applications. Therefore, the efficient cloning and expression of the organic solvent-tolerant lipase is important to its application. In this work, we first found out an organic solvent-tolerant lipase from Staphylococcus saprophyticus M36 and amplified the 741 bp Lipase gene lip3 (GenBank Accession No. FJ979867), by PCR, which encoded a 31.6 kD polypeptide of 247 amino acid residues. But the lipase shared 83% identity with tentative lip3 gene of Staphylococcus saprophyticus (GenBank Accession No. AP008934). We connected the gene with expression vector pET-DsbA, transformed it into Escherichia coli BL21 (DE3), and obtained the recombinant pET-DsbA-lip3. With the induction by 0.4 mmol/L of isopropyl beta-D-thiogalactopyranoside at pH 8.0, OD600 1.0, 25 degrees C for 12 h, the lipase activity reached up to 25.8 U/mL. The lipase expressed was stable in the presence of methanol, n-hexane, and isooctane, n-heptane.
Tang
Yanchong
Y
Enzyme Engineering Laboratory, College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China.
Lu
Yaping
Y
Lü
Fengxia
F
Bie
Xiaomei
X
Guo
Yao
Y
Lu
Zhaoxin
Z
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D008049:Lipase / Q000096:biosynthesis* / Q000235:genetics; D008969:Molecular Sequence Data; D009930:Organic Chemicals / Q000737:chemistry; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D012997:Solvents / Q000737:chemistry*; D057790:Staphylococcus saprophyticus / Q000201:enzymology*
D009930:Organic Chemicals; D011994:Recombinant Proteins; D012997:Solvents; D008049:Lipase