Optimizing a production strategy for a nonspecific nuclease from Yersinia enterocolitica subsp. palearctica in genetically engineered Escherichia coli. Issue 24 (4th December 2019)

• Record Type: Journal Article
• Authors: Ge, Yan; Guo, Senlin; Liu, Tao; Zhao, Chen; Li, Duanhua; Liu, Yangchang; Li, Jinjun; Liang, Tao; Wang, Lu
• Is Part Of: FEMS microbiology letters. Volume 366:Issue 24(2019)
• Publication Date: 2019-12-04
• Subjects: Yersinia enterocolitica -- nonspecific nuclease -- recombinant protein expression -- renaturation; Microbiology -- Periodicals; 579
• DOI: 10.1093/femsle/fnz208
• Languages: English
• ISSNs: 0378-1097

Abstract

ABSTRACT: A nuclease from Yersinia enterocolitica subsp. palearctica (Nucyep) is a newly found thermostable nonspecific nuclease. The heat-resisting ability of this nuclease would be extremely useful in biological research or pharmaceutical production. However, the application of this nuclease is limited because of its poor yield. This research aimed to improve Nucyep productivity by producing a novel genetically engineered Escherichia coli and optimizing the production procedures. After 4 h of induction by lactose, the new genetically engineered E. coli can express a substantial amount of Nucyep in the form of inclusion bodies. The yield was approximately 0.3 g of inclusion bodies in 1 g of bacterial pellets. The inclusion bodies were extracted by sonication and solubilized in an 8 M urea buffer. Protein renaturation was successfully achieved by dilution method. Pure enzyme was obtained after subjecting the protein solution to anion exchange. The Nucyep showed its nonspecific and heat resistant properties as previously reported (Boissinot et al . 2016 ). Through a quantification method, its activity was determined to be 1.3 × 10 6 Kunitz units (K.U.)/mg. These results can serve as a reference for increasing Nucyep production. Abstract : This paper describes an optimised strategy to produce a thermostable nonspecific nuclease in genetically engineered bacteria.