title: Overexpression of PACEsol improves BMP-7 processing in recombinant CHO cells.
abstract: Bone morphogenetic protein-7 (BMP-7) is a member of the TGF-β superfamily and plays a critical role in cartilage, bone, and kidney development. BMP-7 is synthesized as a large precursor and undergoes proteolytic cleavage by subtilisin-like proprotein convertase to secrete the functionally active mature dimer. When CHO cells producing recombinant human BMP-7 (rhBMP) (CHO-BMP-7) were cultivated in a serum-free suspension culture, a significant amount of unwanted precursor forms of rhBMP-7 (ca. 69% of total rhBMP-7), along with the mature form of rhBMP-7, was secreted into the culture medium, likely due to the insufficient amount of the proteolytic cleaving enzyme within the secretory pathway. In order to solve this problem, a soluble form of the paired basic amino acid cleaving enzyme (PACEsol), responsible for the majority of the processing events occurring in the constitutive secretory pathway in mammalian cells, was overexpressed in CHO-BMP-7 cells. Overexpression of PACEsol was effective in processing the precursor forms of BMP-7, while it did not significantly affect cell growth. As a result, the culture supernatants of CHO-BMP-7 cells overexpressing PACEsol contained almost 100% of the mature BMP-7 form. Taken together, the results show that PACEsol overexpression in CHO-BMP-7 cells is an efficient means of increasing the production of mature BMP-7 and facilitating the downstream purification steps by eliminating the need to remove the precursor forms.
authors: Department of Biological Sciences and Graduate School of Nanoscience & Technology, KAIST, Daejeon, Republic of Korea.
authors: Madhavi
authors: M
authors: Sathyamurthy
authors: Jae Seong
authors: JS
authors: Lee
authors: Jin Hyoung
authors: JH
authors: Park
authors: Yeon Jung
authors: YJ
authors: Kim
authors: Ji Yeon
authors: JY
authors: Jeong
authors: Ju Woong
authors: JW
authors: Jang
authors: Gyun Min
authors: GM
authors: Lee
mesh_terms: D000818:Animals; D001709:Biotechnology; D015153:Blotting, Western; D055419:Bone Morphogenetic Protein 7 / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D016466:CHO Cells; D048708:Cell Growth Processes / Q000502:physiology; D002470:Cell Survival / Q000502:physiology; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D045683:Furin / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*
chemical_list: C525819:BMP7 protein, human; D055419:Bone Morphogenetic Protein 7; D011994:Recombinant Proteins; D045683:Furin