Physicochemical properties of recombinant hepatitis B surface antigen expressed in mammalian cell (C127).
The physicochemical properties of recombinant hepatitis B surface antigen (r-HBsAg), which was expressed in C127 mammalian cell were studied. Using roller bottle culture in DMEM supplemented with fetal bovine serum, 10-15 mg/L of r-HBsAg was produced with about 31% of purification yield. The purity of r-HBsAg by HPLC was 99.8% and electron microscopic examination showed homogeneous spherical particle with 22 nm in diameter, a morphological characteristic of HBsAg. The density of r-HBsAg by CsCl density gradient method was 1.19 g/ml and the isoelectric point by Mono P HR 5/20 column was 4.6. The analysis of subunit protein pattern using SDS-PAGE followed by scanning densitometry gave 81.3% of S protein and 18.7% of pre-S protein. Fluorophore-assisted-carbohydrate-electrophoresis analysis showed the relative amount of carbohydrate to protein was 1.7% and its major component was N-acetyl glucosamine, which was about 39% of total carbohydrate. The relative amount of lipid to protein determined by vanillin phosphoric acid method was 32.5% and its major component was phospholipid, which was about 70% of total lipid. The physicochemical properties of C127 mammalian cell-derived r-HBsAg are similar to those of p-HBsAg, suggesting that the r-HBsAg can be used in developing a new preventive vaccine against hepatitis B.
Lee
Y S
YS
College of Pharmacy, Seoul National University, Korea.
Kim
B K
BK
Choi
E C
EC
D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D002241:Carbohydrates / Q000032:analysis; D002460:Cell Line; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002784:Cholesterol / Q000032:analysis; D006224:Cricetinae; D005347:Fibroblasts / Q000378:metabolism; D006514:Hepatitis B Surface Antigens / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification; D007525:Isoelectric Focusing; D008055:Lipids / Q000032:analysis; D051379:Mice; D008854:Microscopy, Electron; D009005:Monosaccharides / Q000032:analysis; D010743:Phospholipids / Q000032:analysis; D011994:Recombinant Proteins / Q000737:chemistry; D014280:Triglycerides / Q000032:analysis
D002241:Carbohydrates; D006514:Hepatitis B Surface Antigens; D008055:Lipids; D009005:Monosaccharides; D010743:Phospholipids; D011994:Recombinant Proteins; D014280:Triglycerides; D002784:Cholesterol