Expression and characterization of a functional human immunodeficiency virus envelope glycoprotein in insect cells.
Recombinant baculoviruses were used to express the gp160 envelope glycoprotein of the human immunodeficiency virus type 1 (HIV-1) and a truncated variant designated gp160(t) which lacks a transmembrane domain. Glycosylation, proteolytic cleavage, secretion, and biological activities of gp160 and gp160(t) 160(t) were studied in Spodoptera frugiperda cells. Both proteins were rapidly glycosylated and initially were found to be totally endo-beta-N-acetyl-D-glucosaminidase H (endo-H) sensitive. However, partial resistance to endo-H was gradually acquired by both molecules. gp160 was found to remain cell-associated, whereas gp160(t) was secreted into the culture medium in large amounts. A fraction of gp160 and gp160(t) appeared to be proteolytically cleaved, and a cleavage product corresponding in size to gp120 was identified in the culture medium. gp160(t) was found to interact specifically with CD4 receptors without any requirement for proteolytic cleavage. The gp160 protein was shown to be expressed on the surface of S. frugiperda cells by direct immunofluorescence. These surface molecules were biologically active, as demonstrated by their ability to induce syncytium formation when cocultivated with HeLa T4 cells.
Wells
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Department of Microbiology, University of Alabama, Birmingham 35294.
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D000118:Acetylglucosaminidase / Q000378:metabolism; D000818:Animals; D015704:CD4 Antigens / Q000378:metabolism; D002460:Cell Line; D015967:Gene Expression Regulation, Viral; D015686:Gene Products, env / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D005822:Genetic Vectors; D006031:Glycosylation; D015699:HIV Envelope Protein gp120 / Q000032:analysis / Q000378:metabolism*; D019302:HIV Envelope Protein gp160; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007304:Insect Viruses / Q000235:genetics; D007700:Kinetics; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D009036:Moths; D011233:Precipitin Tests; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
D015704:CD4 Antigens; D015686:Gene Products, env; D015699:HIV Envelope Protein gp120; D019302:HIV Envelope Protein gp160; D011498:Protein Precursors; D000118:Acetylglucosaminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase