The E3-14.5K integral membrane protein of adenovirus that is required for down-regulation of the EGF receptor and for prevention of TNF cytolysis is O-glycosylated but not N-glycosylated.
The adenovirus E3-14.5K protein is a cytoplasmic integral membrane protein that functions in concert with the E3-10.4K protein to down-regulate the epidermal growth factor receptor and to prevent tumor necrosis factor cytolysis in adenovirus-infected cells. The 14.5K protein migrates as multiple bands in SDS-PAGE, indicating that it undergoes post-translational modification. The 14.5K protein is known to be phosphorylated on serine. We show here that 14.5K can be metabolically labeled with [3H]glucosamine, that the label is labile to alkali, and that the SDS-PAGE band pattern is simplified in a cell line that is defective in O-glycosylation. Thus, 14.5K is O-glycosylated, probably at a single site in the NH2-terminal lumenal domain. The protein was not metabolically labeled with [3H]mannose, and its SDS-PAGE band pattern was not affected by tunicamycin treatment in vivo or endo F treatment in vitro; thus, 14.5K is not N-glycosylated. There was no evidence that the 10.4K protein is glycosylated, and the 10.4K protein was not required for glycosylation of 14.5K. Virtually all 14.5K molecules appear to contain the core disaccharide Gal beta 1-3GalNAc alpha 1-Ser/Thr which is commonly found on mucin-type O-glycoproteins, and neuraminidase digestion experiments indicated that this disaccharide contains terminal sialic acid.
Krajcsi
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P
Institute for Molecular Virology, St. Louis University School of Medicine, Missouri 63110.
Tollefson
A E
AE
Wold
W S
WS
D017456:Adenovirus Early Proteins; D000260:Adenoviruses, Human / Q000737:chemistry*; D000595:Amino Acid Sequence; D002240:Carbohydrate Sequence; D002478:Cells, Cultured; D015536:Down-Regulation; D066246:ErbB Receptors / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D066298:In Vitro Techniques; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D009856:Oncogene Proteins, Viral / Q000378:metabolism*; D011499:Protein Processing, Post-Translational
D017456:Adenovirus Early Proteins; D008562:Membrane Glycoproteins; D009856:Oncogene Proteins, Viral; D066246:ErbB Receptors