Role of carbohydrate processing and calnexin binding in the folding and activity of the HN protein of Newcastle disease virus.
The role of carbohydrate processing and calnexin binding in the folding pathway and activity of the hemagglutinin-neuraminidase (HN) protein of Newcastle disease virus (NDV) was explored in infected cells using the inhibitor castanospermine (CST). Calnexin-HN protein complexes were demonstrated by coimmunoprecipitation using antibody specific for calnexin or HN protein. As in other systems, this complex was not detected in CST treated cells. In cells incubated in CST, the synthesis and stability of the HN protein was unaffected. However, as monitored by the appearance of conformationally sensitive antigenic sites, the folding of the HN protein in CST treated cells was approximately twice as slow than in untreated cells. This folding was ultimately efficient since there was no evidence for significant amounts of irreversibly aggregated forms which never acquired a mature conformation. Most significantly, the folding sequence as measured by the order of appearance of conformationally sensitive antigenic sites (McGinnes and Morrison, Virology 199, 255) was unaffected by CST. Thus while calnexin functions to speed the folding of the HN protein, it is not required for the folding of this protein. In addition, the protein synthesized in the presence of CST had significant levels of neuraminidase and hemagglutination activity suggesting that processing of the carbohydrate has a minimal role in the activity of the protein.
McGinnes
L W
LW
Department of Molecular Genetics and Microbiology, University of Massachusetts Medical School, Worcester 01655, USA.
Morrison
T G
TG
D000818:Animals; D000956:Antigens, Viral / Q000378:metabolism; D001692:Biological Transport; D019556:COS Cells; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D037281:Calnexin; D050260:Carbohydrate Metabolism; D002462:Cell Membrane; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015707:HN Protein / Q000737:chemistry / Q000378:metabolism*; D007212:Indolizines / Q000494:pharmacology; D007700:Kinetics; D009522:Newcastle disease virus / Q000378:metabolism*; D011487:Protein Conformation; D017510:Protein Folding; D054836:Indolizidines
D000956:Antigens, Viral; D002135:Calcium-Binding Proteins; D037281:Calnexin; D004791:Enzyme Inhibitors; D015707:HN Protein; D007212:Indolizines; C037806:castanospermine; D054836:Indolizidines