Overproduction of recombinant human hepatocyte growth factor in Chinese hamster ovary cells.
Hepatocyte growth factor (HGF) is a potent multi-functional protein that affects morphogenesis, cell migration, organ regeneration, and tumor invasion in various tissues, and has thus been considered to have potential as a therapeutic target in various diseases. In our current study, we established Chinese hamster ovary (CHO) cells overexpressing recombinant human HGF (rhHGF) protein and in a 5 day batch culture process using a 7.5l bioreactor (5l working volume) and serum-free medium these cells could produce over 13 mg/l of rhHGF protein. The recombinant protein was then purified to homogeneity from the culture supernatant using a two-step chromatographic procedure that resulted in about a 35% recovery rate. This purified rhHGF was found to be a mixture of inactive pro-HGF and an active heterodimeric form of this protein with a higher molecular weight than its counterpart expressed from insect cells. This finding suggests that the glycosylation of rhHGF protein in CHO cells differs from that in insect cells. Inactive pro-HGF was found to rapidly convert to the active heterodimeric form of HGF in the presence of FBS (Fetal Bovine Serum), suggesting that this process would occur also when injected into human body. We further demonstrate in cell proliferation and scattering activity assays that our purified rhHGF protein preparation is functionally active with a half-maximal effective concentration of 36.3 ng/ml.
Park
Jeong Soo
JS
Neurotech Pharmaceuticals Co. Ltd., Suwon 443-766, Republic of Korea.
Kim
Hyunjoo
H
Park
Junho
J
Yu
Sungryul
S
Kim
Dongjun
D
Lee
Jongmin
J
Oh
Hankyu
H
Baek
Kwanghee
K
Yoon
Jaeseung
J
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D017228:Hepatocyte Growth Factor / Q000096:biosynthesis*; D011994:Recombinant Proteins / Q000096:biosynthesis*
C066855:HGF protein, human; D011994:Recombinant Proteins; D017228:Hepatocyte Growth Factor