High-level expression and characterization of hepatitis B virus surface antigen in silkworm using a baculovirus vector.
The Bombyx mori nuclear polyhedrosis virus (BmNPV) in the silkworm was used successfully for mass production of biologically active foreign genes under the control of the polyhedrin promoter. This system was adapted for the production of large amounts of hepatitis B virus surface antigens (HBsAg). The DNA fragments coded for the middle protein, which is composed of the S protein with the pre-S2 region, were cloned, the signal protein gene of beta-IFN was added, and both were inserted into a cloning vector. After co-transfection with wild-type BmNPV, stable recombinant viruses were isolated by the limiting dilution method. Infected silkworm larvae with the recombinants expressed HBsAg at high levels (400-600 micrograms/ml). These products, consisting of two polypeptides with molecular weights of approximately 25,000 (p25) and glycosylated P25 (GP28), were purified as assembled 22-nm particles. We demonstrated that HBsAg from silkworms consists of S protein with 7 amino acids of Pre-S2.
Higashihashi
N
N
Molecular Biology Research Laboratory, Daiichi Pharmaceutical Co., Ltd., Edogawa-ku, Tokyo, Japan.
Arai
Y
Y
Enjo
T
T
Horiuchi
T
T
Saeki
Y
Y
Sakano
K
K
Sato
Y
Y
Takeda
K
K
Takashina
S
S
Takahashi
T
T
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D001483:Base Sequence; D015153:Blotting, Western; D012831:Bombyx; D004247:DNA; D005822:Genetic Vectors; D006514:Hepatitis B Surface Antigens / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D008969:Molecular Sequence Data; D009841:Oligonucleotides; D010957:Plasmids; D014162:Transfection
D006514:Hepatitis B Surface Antigens; D009841:Oligonucleotides; D004247:DNA