title: Development of a new platform for secretory production of recombinant proteins in Corynebacterium glutamicum.
abstract: Corynebacterium glutamicum, which has been for long an industrial producer of various L-amino acids, nucleic acids, and vitamins, is now also regarded as a potential host for the secretory production of recombinant proteins. To harness its potential as an industrial platform for recombinant protein production, the development of an efficient secretion system is necessary. Particularly, regarding protein production in large-scale bioreactors, it would be appropriate to develop a secretory expression system that is specialized for high cell density cultivation conditions. Here we isolated a new signal peptide that mediates the efficient secretion of recombinant proteins under high cell density cultivation conditions. The secretome of C. glutamicum ATCC 13032 under high cell density cultivation conditions was initially investigated, and one major protein was identified as a hypothetical protein encoded by cg1514. Novel secretory production systems were then developed using the Cg1514 signal peptide and its own promoter. Efficient protein secretion was demonstrated using three protein models: endoxylanase, α-amylase, and camelid antibody fragment (VHH). For large-scale production, fed-batch cultivations were also conducted and high yields were successfully achieved--as high as 1.07 g/L (endoxylanase), 782.6 mg/L (α-amylase), and 1.57 g/L (VHH)--in the extracellular medium. From the culture media, all model proteins could be simply purified by one-step column chromatography with high purities and recovery yields. To the best of our knowledge, this is the first report of the development of an efficient secretory expression system by secretome analysis under high cell density cultivation conditions in C. glutamicum.
authors.affiliation: Department of Chemical and Biomolecular Engineering, BK21 Plus program, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea.
authors.forename: Sung Sun
authors.initials: SS
authors.lastname: Yim
authors.affiliation: Department of Chemical and Biomolecular Engineering, BK21 Plus program, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea.
authors.forename: Jae Woong
authors.initials: JW
authors.lastname: Choi
authors.affiliation: Department of Chemical and Biomolecular Engineering, BK21 Plus program, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea.
authors.forename: Roo Jin
authors.initials: RJ
authors.lastname: Lee
authors.affiliation: Department of Chemical and Biomolecular Engineering, BK21 Plus program, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea.
authors.forename: Yong Jae
authors.initials: YJ
authors.lastname: Lee
authors.affiliation: Department of Chemical and Biomolecular Engineering, BK21 Plus program, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea.
authors.forename: Se Hwa
authors.initials: SH
authors.lastname: Lee
authors.affiliation: Bio R&D Center, CJ CheilJedang, 92 Gayang-dong, Gangseo-gu, Seoul, 175-724, Republic of Korea.
authors.forename: So Young
authors.initials: SY
authors.lastname: Kim
authors.affiliation: Department of Chemical and Biomolecular Engineering, BK21 Plus program, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea. kjjeong@kaist.ac.kr.|Institute for the BioCentury, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea. kjjeong@kaist.ac.kr.
authors.forename: Ki Jun
authors.initials: KJ
authors.lastname: Jeong
mesh_terms: D019149:Bioreactors / Q000382:microbiology; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D043364:Endo-1,4-beta Xylanases / Q000235:genetics / Q000378:metabolism; D060847:Metabolic Engineering / Q000379:methods*; D021382:Protein Sorting Signals; D011994:Recombinant Proteins / Q000235:genetics* / Q000378:metabolism*; D061905:Single-Domain Antibodies / Q000235:genetics / Q000378:metabolism; D000516:alpha-Amylases / Q000235:genetics / Q000378:metabolism
chemical_list: D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D061905:Single-Domain Antibodies; D000516:alpha-Amylases; D043364:Endo-1,4-beta Xylanases
keywords: Cg1514; Corynebacterium glutamicum; fed-batch cultivation; secretion; secretome