Biochemical and immunological characterization of the bovine leukemia virus (BLV) envelope glycoprotein (gp51) produced in Saccharomyces cerevisiae.
The nucleotide sequence coding for bovine leukemia virus (BLV) envelope glycoprotein gp51 was inserted into a yeast-Escherichia coli shuttle vector carrying the promoter and secretion signal sequence of PHO5 (the yeast gene coding for repressible acid phosphatase) and the CYC1 transcriptional terminator. Yeast cells transformed by this construction synthesized gp51 after PHO5 induction by inorganic phosphate deprivation. The yeast-expressed gp51 was partially glycosylated into heterodisperse protein molecules ranging from 40 to 48 kDa. No gp51 was excreted in the culture medium. The amount of protein accumulated in yeast cells was estimated to reach 0.06% of soluble proteins. This modest level of expression seemed to be due to the toxicity of gp51 to the yeast cell. The yeast-expressed gp51 products were used in enzyme-linked immunosorbent assays for the detection of antibodies in sera from BLV-infected animals; they were also screened for the presence of well-defined biological epitopes. In both studies poor reactivity was observed. Rabbits immunized with the recombinant gp51 showed high antibody titers to native BLV gp51. However, these antibodies did not neutralize BLV in vitro.
Legrain
M
M
Unit of Microbiology, Faculty of Agronomy, Gembloux, Belgium.
Portetelle
D
D
Dumont
J
J
Burny
A
A
Hilger
F
F
D000135:Acid Phosphatase / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology; D003001:Cloning, Molecular; D003603:Cytotoxins / Q000096:biosynthesis; D004279:DNA, Viral / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D006023:Glycoproteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000633:toxicity; D006031:Glycosylation; D001909:Leukemia Virus, Bovine / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals / Q000235:genetics; D012190:Retroviridae / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D014759:Viral Envelope Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000633:toxicity
D000914:Antibodies, Viral; D000956:Antigens, Viral; D003603:Cytotoxins; D004279:DNA, Viral; D006023:Glycoproteins; D021382:Protein Sorting Signals; D014759:Viral Envelope Proteins; D000135:Acid Phosphatase