Translation and membrane insertion of the hemagglutinin-neuraminidase glycoprotein of Newcastle disease virus.
The hemagglutinin-neuraminidase (HN) protein of paramyxoviruses is likely in the unusual class of glycoproteins with the amino terminus cytoplasmic and the carboxy terminus lumenal or external to the cell. The properties of the membrane insertion of the HN protein of Newcastle disease virus, a prototype paramyxovirus, were explored in wheat germ extracts containing microsomal membranes. HN protein was inserted into membranes cotranslationally, resulting in a glycosylated protein completely resistant to trypsin and proteinase K digestion. No detectable posttranslation insertion occurred. Insertion required signal recognition particle. Signal recognition particle in the absence of membranes inhibited HN protein synthesis. Comparisons of the trypsin digestion products of the HN protein made in the cell-free system with newly synthesized HN protein from infected cells showed that the cell-free product was in a conformation different from that of the pulse-labeled protein in infected cells. First, trypsin digestion of intact membranes from infected cells reduced the size of the 74,000-dalton HN protein by approximately 1,000 daltons, whereas trypsin digestion of HN protein made in the cell-free system had no effect on the size of the protein. Second, trypsin digestion of Triton X-100-permeabilized membranes isolated from infected cells resulted in a 67,000-dalton trypsin resistant HN protein fragment. A trypsin-resistant core of comparable size was not present in the digestion products of in-vitro-synthesized HN protein. Evidence is presented that the newly synthesized HN protein in infected cels contain intramolecular disulfide bonds not present in the cell-free product.
D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D015707:HN Protein; D006389:Hemagglutinins, Viral / Q000235:genetics*; D007425:Intracellular Membranes / Q000378:metabolism; D008861:Microsomes / Q000378:metabolism; D009522:Newcastle disease virus / Q000235:genetics* / Q000378:metabolism; D010944:Plants / Q000378:metabolism; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D014158:Transcription, Genetic; D014908:Triticum / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000378:metabolism
D015707:HN Protein; D006389:Hemagglutinins, Viral; D014759:Viral Envelope Proteins