title: A novel cold-adapted lipase from Sorangium cellulosum strain So0157-2: gene cloning, expression, and enzymatic characterization.
abstract: Genome sequencing of cellulolytic myxobacterium Sorangium cellulosum reveals many open-reading frames (ORFs) encoding various degradation enzymes with low sequence similarity to those reported, but none of them has been characterized. In this paper, a predicted lipase gene (lipA) was cloned from S. cellulosum strain So0157-2 and characterized. lipA is 981-bp in size, encoding a polypeptide of 326 amino acids that contains the pentapeptide (GHSMG) and catalytic triad residues (Ser114, Asp250 and His284). Searching in the GenBank database shows that the LipA protein has only the 30% maximal identity to a human monoglyceride lipase. The novel lipA gene was expressed in Escherichia coli BL21 and the recombinant protein (r-LipA) was purified using Ni-NTA affinity chromatography. The enzyme hydrolyzed the p-nitrophenyl (pNP) esters of short or medium chain fatty acids (≤C(10)), and the maximal activity was on pNP acetate. The r- LipA is a cold-adapted lipase, with high enzymatic activity in a wide range of temperature and pH values. At 4 °C and 30 °C, the K(m) values of r-LipA on pNP acetate are 0.037 ± 0.001 and 0.174 ± 0.006 mM, respectively. Higher pH and temperature conditions promoted hydrolytic activity toward the pNP esters with longer chain fatty acids. Remarkably, this lipase retained much of its activity in the presence of commercial detergents and organic solvents. The results suggest that the r-LipA protein has some new characteristics potentially promising for industrial applications and S. cellulosum is an intriguing resource for lipase screening.
authors.affiliation: State Key Laboratory of Microbial Technology, School of Life Science, Shandong University, Jinan 250100, China; E-Mails:  cyysdu@gmail.com  (Y.-Y.C.);  qyk-1@163.com  (Y.-K.Q.);  lizhifeng@sdu.edu.cn  (Z.-F.L.);  wuzhihong@sdu.edu.cn  (Z.-H.W.);  liuhong05@sdu.edu.cn  (H.L.).
authors.forename: Yuan-Yuan
authors.initials: YY
authors.lastname: Cheng
authors.forename: Yun-Kai
authors.initials: YK
authors.lastname: Qian
authors.forename: Zhi-Feng
authors.initials: ZF
authors.lastname: Li
authors.forename: Zhi-Hong
authors.initials: ZH
authors.lastname: Wu
authors.forename: Hong
authors.initials: H
authors.lastname: Liu
authors.forename: Yue-Zhong
authors.initials: YZ
authors.lastname: Li
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003080:Cold Temperature; D030541:Databases, Genetic; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008049:Lipase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D008994:Monoacylglycerol Lipases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009231:Myxococcales / Q000201:enzymology*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D008994:Monoacylglycerol Lipases; D008049:Lipase
references.citation: Biotechnol Bioeng. 1987 Jul;30(1):81-7
references.pmid: 18576586
references.citation: J Biosci Bioeng. 2001;92(2):144-8
references.pmid: 16233074
references.citation: J Microbiol Biotechnol. 2009 Feb;19(2):128-35
references.pmid: 19307760
references.citation: Appl Environ Microbiol. 1998 Feb;64(2):486-91
references.pmid: 9464382
references.citation: Int J Mol Sci. 2011;12(6):3950-65
references.pmid: 21747717
references.citation: Chirality. 1996;8(7):481-89
references.pmid: 8970745
references.citation: J Biotechnol. 2003 Dec 19;106(2-3):233-53
references.pmid: 14651865
references.citation: Curr Opin Chem Biol. 2002 Apr;6(2):145-50
references.pmid: 12038997
references.citation: Nat Biotechnol. 2007 Nov;25(11):1281-9
references.pmid: 17965706
references.citation: Eur J Biochem. 1992 Jan 15;203(1-2):25-32
references.pmid: 1730231
references.citation: Annu Rev Microbiol. 1999;53:315-51
references.pmid: 10547694
references.citation: Biotechnol Appl Biochem. 1999 Apr;29(2):119-31
references.pmid: 10075908
references.citation: Proc Natl Acad Sci U S A. 1985 May;82(10):3192-6
references.pmid: 3858815
references.citation: Angew Chem Int Ed Engl. 1998 Jul 3;37(12):1608-1633
references.pmid: 29711530
references.citation: Microbiol Rev. 1990 Dec;54(4):473-501
references.pmid: 1708086
references.citation: J Microbiol Biotechnol. 2007 Apr;17(4):604-10
references.pmid: 18051271
references.citation: Protein Expr Purif. 2006 Apr;46(2):309-15
references.pmid: 16169745
references.citation: Biotechnol Adv. 2008 Sep-Oct;26(5):457-70
references.pmid: 18571355
references.citation: Lett Appl Microbiol. 2003;37(4):354-9
references.pmid: 12969503
references.citation: Ann Clin Biochem. 2001 Jul;38(Pt 4):365-70
references.pmid: 11471878
references.citation: Nucleic Acids Res. 2008 Jan;36(Database issue):D281-8
references.pmid: 18039703
references.citation: Protein Expr Purif. 2004 Mar;34(1):103-10
references.pmid: 14766305
references.citation: Appl Microbiol Biotechnol. 2006 Apr;70(3):321-6
references.pmid: 16088345
references.citation: J Microbiol. 2010 Aug;48(4):438-44
references.pmid: 20799084
references.citation: J Mol Biol. 2010 Feb 26;396(3):663-73
references.pmid: 19962385
references.citation: Mol Biol Rep. 2009 Jul;36(6):1515-9
references.pmid: 18773306
references.citation: Appl Environ Microbiol. 1991 May;57(5):1560-2
references.pmid: 16348494
references.citation: Appl Microbiol Biotechnol. 2011 May;90(3):971-80
references.pmid: 21336927
references.citation: Biochem J. 1999 Oct 1;343 Pt 1:177-83
references.pmid: 10493927
references.citation: Appl Microbiol Biotechnol. 2009 Jan;81(5):865-74
references.pmid: 18773201
references.citation: Mar Biotechnol (NY). 2008 Sep-Oct;10(5):612-21
references.pmid: 18461394