Development of a transposable elements free escherichia coli for the efficient production of recombinant therapeutic proteins = 의약용 재조합 단백질의 효율적인 생산을 위한 전이성 유전인자 제거 대장균 개발 연구

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Genome stability and integrity of a host strain are critical for the production of high quali-ty bioproducts from industrial microorganisms. However, the genomes of microorgan-isms are littered with transposable elements (TEs) called “jumping genes”, which are re-combinogenic factor causing genome instability and genetic variations. These elements cause many genetic rearrangements such as inversion, deletion, duplication and transposi-tion, thus causing adverse effects such as production of unwanted by-products and non-homogeneous products and masking expression of genes. To minimize these harmful ef-fects of TEs of Escherichia coli, we developed a TEs-free strain, in which all of TEs in-cluding insertion sequences (IS), transposons, phages and cryptic phages, were removed from E. coli MG1655 genome by a rapid markerless deletion system. Here, we compared the expression profiles of recombinant protein such as tumor necrosis factor-related apop-tosis-inducing ligand (TRAIL) and bone morphogenetic protein-2 (BMP2) in MG1655 and TEs-free strain. The hopping of ISs (IS1, IS3 or IS5) into the TRAIL and BMP2 genes oc-curred at the rate of $~10^{-8}/gene/h$ in MG1655 whereas no IS hopping was observed in TEs-free strain. Even though IS hopping occurred very rarely $(10^{-8} /gene/h)$, the IS-inserted TRAIL and BMP2 clones became dominant (~52 % of total population) in 28 h after fer-mentation because of its rapid growth, significantly reducing production of recombinant therapeutic proteins in batch fermentation. Our observation clearly indicates that TEs jumping is detrimental to the production of recombinant enzymes, strongly emphasizing the necessity of developing a TEs-free host strain. Thus, our TEs-free strain will be use-ful as a robust and suitable host for the production of recombinant therapeutic proteins.
Kim, Sun-Changresearcher김선창
한국과학기술원 : 생명과학과,
Issue Date
591771/325007  / 020078013

학위논문(박사) - 한국과학기술원 : 생명과학과, 2014.8, [ v, 103p ]


transposable element; 인서션 시퀀스; 전이성 유전인자 제거 대장균; 전이성 유전인자; IS-hopping; Insertion sequence; jumping genes; TEs-free E. coli

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