High level expression and characterization of a novel thermostable, organic solvent tolerant, 1,3-regioselective lipase from Geobacillus sp. strain ARM
ABSTRACT The mature ARM lipase gene was cloned into the pTrcHis expression vector and over-expressed in Escherichia coli TOP10 host. The optimum lipase expression was obtained after 18 h post induction incubation with 1.0mM IPTG, where the lipase activity was approximately 1623-fold higher than wild type. A rapid, high efficient, one-step purification of the His-tagged recombinant lipase was achieved using immobilized metal affinity chromatography with 63.2% recovery and purification factor of 14.6. The purified lipase was characterized as a high active (7092 U mg(-1)), serine-hydrolase, thermostable, organic solvent tolerant, 1,3-specific lipase with a molecular weight of about 44 kDa. The enzyme was a monomer with disulfide bond(s) in its structure, but was not a metalloenzyme. ARM lipase was active in a broad range of temperature and pH with optimum lipolytic activity at pH 8.0 and 65°C. The enzyme retained 50% residual activity at pH 6.0-7.0, 50°C for more than 150 min.
Full-textDOI: · Available from: Afshin Ebrahimpour, May 16, 2014
- SourceAvailable from: Aleksey S Rozanov
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- "Many genes of thermophilic enzymes have been isolated in recent years from the genomes of Geobacillus for their further heterological expression in mesophylic bacteria. Among these enzymes, lipases are found (AbdellFatt tah, Gaballa, 2008; QuintanaaCastro et al., 2009; Cheong et al., 2011; Ebrahimpour et al., 2011; Balan et al., 2012). They take part in the decomposition of lignocellulosic biomass: βglucosidase (Shallom et al., RUSSIAN JOURNAL OF GENETICS: APPLIED RESEARCH Vol. 4 No. 3 2014 2005; BenDavid et al., 2007; Wagschal et al., 2009; Ratnadewi et al., 2013), endoglucanase (Ng et al., 2009), xylanase (Wu et al., 2006; Canakci et al., 2007, 2012; Gerasimova, Kuisiene, 2012; Liu et al., 2012; Verma et al., 2013), and many other proteins. "
ABSTRACT: Thermophilic bacteria are being extensively used in biotechnology. Species of the genus Geobacillus rank among the most prospective thermophiles. The current methods of genetic and metabolic engineering of these microorganisms are considered in the article. Examples of their use in various branches of biotechnology are presented.05/2014; 4(3):218-226. DOI:10.1134/S2079059714030046
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ABSTRACT: Paired surface transverse wave resonators operating near 500 MHz are being used as the sensing element in a chemical/biological detector capable of operating in water. Initially, the design of the surface transverse wave detector is discussed. Then, the major problems uncovered with the prototype detector are reviewed with an emphasis on solutions to these problems that will be implemented in a second-generation “smart” detectorFrequency Control Symposium, 1994. 48th., Proceedings of the 1994 IEEE International; 07/1994
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ABSTRACT: The Rhizopus oryzae lipase containing prosequence was expressed in Pichia pastoris. Recombinant lipase subunit showed a molecular mass of 32 kDa. The maximum activity of recombinant lipase obtained from Mut(s) recombinant was 90 IU/ml. The enzyme was stable in broad ranges of temperatures and pH, with the optimal temperature at 35 °C and pH 7.0. The crude recombinant R. oryzae lipase can be directly used for the transesterification of plant oils at high-water content of 60-100% (w/w) based on oil weight. The addition of 80% water to the transesterification systems resulted in the yield of methyl ester of 95%, 94% and 92% after 72 h using soybean oil, Jatropha curcas seed raw oil and Pistacia chinensis seed raw oil as raw material, respectively. These results indicate that the recombinant lipase is an effective biocatalyst for enzymatic biodiesel production.Bioresource Technology 07/2011; 102(20):9810-3. DOI:10.1016/j.biortech.2011.07.070 · 5.04 Impact Factor