title: Enhanced production of recombinant proteins with Corynebacterium glutamicum by deletion of insertion sequences (IS elements).
abstract: BACKGROUND
  In most bacteria, various jumping genetic elements including insertion sequences elements (IS elements) cause a variety of genetic rearrangements resulting in harmful effects such as genome and recombinant plasmid instability. The genetic stability of a plasmid in a host is critical for high-level production of recombinant proteins, and in this regard, the development of an IS element-free strain could be a useful strategy for the enhanced production of recombinant proteins. Corynebacterium glutamicum, which is a workhorse in the industrial-scale production of various biomolecules including recombinant proteins, also has several IS elements, and it is necessary to identify the critical IS elements and to develop IS element deleted strain.
  
  
  RESULTS
  From the cultivation of C. glutamicum harboring a plasmid for green fluorescent protein (GFP) gene expression, non-fluorescent clones were isolated by FACS (fluorescent activated cell sorting). All the isolated clones had insertions of IS elements in the GFP coding region, and two major IS elements (ISCg1 and ISCg2 families) were identified. By co-cultivating cells harboring either the isolated IS element-inserted plasmid or intact plasmid, it was clearly confirmed that cells harboring the IS element-inserted plasmids became dominant during the cultivation due to their growth advantage over cells containing intact plasmids, which can cause a significant reduction in recombinant protein production during cultivation. To minimize the harmful effects of IS elements on the expression of heterologous genes in C. glutamicum, two IS element free C. glutamicum strains were developed in which each major IS element was deleted, and enhanced productivity in the engineered C. glutamicum strain was successfully demonstrated with three models: GFP, poly(3-hydroxybutyrate) [P(3HB)] and γ-aminobutyrate (GABA).
  
  
  CONCLUSIONS
  Our findings clearly indicate that the hopping of IS elements could be detrimental to the production of recombinant proteins in C. glutamicum, emphasizing the importance of developing IS element free host strains.
authors.affiliation: Department of Chemical and Biomolecular Engineering (BK Plus program), KAIST, 291 Daehakro, Yuseong-gu, Daejeon, 34141, Republic of Korea. jwoongci@gmail.com.
authors.forename: Jae Woong
authors.initials: JW
authors.lastname: Choi
authors.affiliation: Department of Chemical and Biomolecular Engineering (BK Plus program), KAIST, 291 Daehakro, Yuseong-gu, Daejeon, 34141, Republic of Korea. vidalsungsun@gmail.com.
authors.forename: Sung Sun
authors.initials: SS
authors.lastname: Yim
authors.affiliation: Department of Chemical and Biomolecular Engineering (BK Plus program), KAIST, 291 Daehakro, Yuseong-gu, Daejeon, 34141, Republic of Korea. mjflorakim@gmail.com.
authors.forename: Min Jeong
authors.initials: MJ
authors.lastname: Kim
authors.affiliation: Department of Chemical and Biomolecular Engineering (BK Plus program), KAIST, 291 Daehakro, Yuseong-gu, Daejeon, 34141, Republic of Korea. kjjeong@kaist.ac.kr.|Institute for the BioCentury, KAIST, 291 Daehakro, Yuseong-gu, Daejeon, 34141, Republic of Korea. kjjeong@kaist.ac.kr.
authors.forename: Ki Jun
authors.initials: KJ
authors.lastname: Jeong
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D048230:Corynebacterium glutamicum / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics*; D017384:Sequence Deletion
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins