Biochemical properties of SV40 large tumor antigen as a glycosylated protein.
The large tumor antigen (T-ag) of SV40 is a virus-encoded polypeptide that provides multiple biological activities required for virus replication and cellular transformation. T-ag is an exceptional model for the study of protein processing, because it displays a variety of chemical modifications and an unusual dual subcellular distribution. The cellular mechanisms responsible for the synthesis and processing of T-ag are unknown. With respect to glycosylation, this has been related to a lack of knowledge of the biochemical properties of T-ag as a glycoprotein. Several such properties are characterized here. We found that T-ag is glycosylated at multiple sites on the polypeptide chain. The oligosaccharides appear to belong to a single size class, molecular weight approximately 400, and the linkage between the polypeptide and the carbohydrate side chain is sensitive to beta-elimination under mild alkaline conditions. At least one glycosylation site was localized to the region between amino acids 1 and 272 (probably between residues 83 and 272), and at least one additional site was localized to a separate region, between amino acids 523 and 708. The results of cycloheximide experiments suggested that glycosylation of T-ag is a cotranslational event, and both the nuclear and the membrane-associated forms of T-ag appeared to be glycosylated. The results of these studies verify previous conclusions that the cellular secretory pathway is not involved in the glycosylation of T-ag; instead, a cytoplasmic mechanism might be involved.
Jarvis
D L
DL
Department of Virology and Epidemiology, Baylor College of Medicine, Houston, Texas 77030.
Butel
J S
JS
D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000032:analysis*; D002236:Carbohydrate Conformation; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D003513:Cycloheximide / Q000494:pharmacology; D005690:Galactose / Q000378:metabolism; D006031:Glycosylation; D008715:Methionine / Q000378:metabolism; D010449:Peptide Mapping; D014176:Protein Biosynthesis
D000952:Antigens, Polyomavirus Transforming; D003513:Cycloheximide; D008715:Methionine; D005690:Galactose