Structure of glycosylated and unglycosylated gag polyproteins of Rauscher murine leukemia virus: carbohydrate attachment sites

Abstract

The structural relationships among the gag polyproteins Pr655"9, Pr75 a8, and gPr80959 of Rauscher murine leukemia virus were studied by endoglycosidase H digestion and formic acid cleavage. Fragments were identified by precipitation with specific antisera to constituent virion structural proteins followed by onedimensional mapping. Endoglycosidase H reduced the size of gPr80959 to that of Pr759'a. By comparing fragments of gPr80g'w and the apoprotein Pr759'9, the former was shown to contain two mannose-rich oligosaccharide units. By comparing fragments of Pr65""9 and Pr755 ¶9, the latter was shown to differ from Pr659"9 at the amino terminus by the presence of a leader peptide approximately 7,000 daltons in size. The internal and carboxyl-terminal peptides of the two unglycosylated polyproteins were not detectably different. The location of the two N-linked carbohydrate chains in gPr80"g" has been specified. One occurs in the carboxyl-terminal half of the polyprotein at asparagine,77 of the p30 sequence and the other is found in a 23,000-dalton fragment located in the amino-terminal region of gPr809a9 and containing the additional amino acid sequences not found in Pr65gag plus a substantial portion of p15. MATERIALS AND METHODS Cells and cell culture. JLS-V9 cells (36) chronically infected with R-MuLV were obtained from the Viral Resources Laboratory (Frederick Cancer Research Center, Frederick, Md.). Fetal calf serum was purchased from GIBCO Laboratories (Grand Island, N.Y.). Eagle minimal essential medium (EMEM) and methionine-free EMEM were obtained from Flow Laboratories (Rockville, Md.). Cells were grown on Corning plastic ware in EMEM supplemented with 10% heat-inactivated fetal calf serum, penicillin (100 IU/ml), and streptomycin (100 ,ug/ 581

The gag gene products of human immunodeficiency virus type 1: alignment within the gag open reading frame, identification of posttranslational modifications, and evidence for alternative gag precursors

“…1A, lane My) that was not am'endable to' direct sequencing. The amino-terminal gag proteins -of certain retroviruses have been shown to be myristoylated (19,43,44 Extracellular virus packaged abundant myristoylated p16 (MA) and much sm'aller quantities of p55 (Fig. 1A, lanes M,My and My).…”

“…Alternative forms of gag proteins have been observed in other retroviral systems. For instance, a glycosylated p85 gag protein, gPr8V'g, is expressed on the surface murine leukemia virus-infected cells (43). It is apparently initiated within the pre-gag region (43,46).…”

“…For instance, a glycosylated p85 gag protein, gPr8V'g, is expressed on the surface murine leukemia virus-infected cells (43). It is apparently initiated within the pre-gag region (43,46). Cells expressing this protein have been shown to serve as targets for cytotoxic T lymphocytes (20, 49).…”

DNA sequence of the viral and cellular src gene of chickens. 1. Complete nucleotide sequence of an EcoRI fragment of recovered avian sarcoma virus which codes for gp37 and pp60src

“…These two gene products were localized as described in the text. Open squares indicate the possible glycosylation sites (32). Closed squares and closed circles indicate the sites for tyrosine phosphorylation (34) and the possible site for serine phosphorylation (21), respectively.…”

“…The sequence of tripeptides, Asn-X-Thr or Asn-X-Ser, which are believed to be the sites for glycosylation (32), can be found at three places, positions 52 to 54, 100 to 102, and 181 to 183.…”