Expression of rat alpha-fetoprotein cDNA in Escherichia coli and in yeast.
Rat alpha-fetoprotein (AFP) cDNA spanning the complete coding region was cloned and expressed in Escherichia coli as well as in yeast, Saccharomyces cerevisiae. The recombinant AFPs (rAFPs) were purified and characterized. The molecular weights determined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis were 65,000 for E. coli rAFP and 69,000 for yeast rAFP. Amino acid and N-terminal sequence analyses indicated that yeast cells produced mature AFP by processing the signal peptide properly but E. coli rAFP lacked the N-terminal 53 amino acid residues of preAFP. The yeast rAFP was found to be indistinguishable from authentic AFP in the Ouchterlony immunodiffusion test, radioimmunoassay and estradiol-binding assay while E. coli rAFP was less reactive in these tests. These observations indicated that the rAFP expressed in yeast emulated the properties of authentic AFP.
Nishi
S
S
Department of Biochemistry, Hokkaido University School of Medicine.
Koyama
Y
Y
Sakamoto
T
T
Soda
M
M
Kairiyama
C B
CB
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D005786:Gene Expression Regulation; D008969:Molecular Sequence Data; D008970:Molecular Weight; D021382:Protein Sorting Signals / Q000235:genetics; D051381:Rats; D012441:Saccharomyces cerevisiae / Q000235:genetics; D000509:alpha-Fetoproteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification
D021382:Protein Sorting Signals; D000509:alpha-Fetoproteins; D004247:DNA