[Cloning and expression of rat alpha-fetoprotein cDNA in Escherichia coli].
AFP is a major serum protein during ontogeny and is synthesized mainly by the mammalian fetal liver and yolk sac. Its synthesis ceases early in postnatal life and its reappearance in the serum in adult is a sign of hepatoma or yolk sac tumor, since these tumors produce AFP. This paper describes the cloning of rat AFP cDNA spanning complete coding region, its expression in E. coli and characterization of this recombinant AFP. The determination of the nucleotide sequence and cell-free translation of purified AFPmRNA suggested that rat AFP was synthesized as a precursor with a signal peptide of 24 amino acids followed by mature AFP of 587 amino acids. An expression vector was constructed with the cDNA and the introduction of the plasmid into E. coli resulted in the production of immunologically reactive AFP with a molecular weight of 65,000. The recombinant AFP was highly purified by immunoaffinity chromatography followed by SDS-PAGE. Analysis of the amino acids sequence indicated that the product was AFP lacking N-terminal 53 amino acid residues of preAFP.
Soda
M
M
Department of Biochemistry, Hokkaido University School of Medicine, Sapporo, Japan.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000235:genetics*; D008969:Molecular Sequence Data; D051381:Rats; D000509:alpha-Fetoproteins / Q000235:genetics*
D004269:DNA, Bacterial; D000509:alpha-Fetoproteins