Secretory protein translocation in a neurospora crassa in vitro system. Hydrolysis of a nucleoside triphosphate is required for posttranslational translocation.
An in vitro translocation system has been reconstituted with subcellular fractions from the cell wall-less mutant of Neurospora crassa (fz;sg;os-1). Prepro alpha factor and invertase, secretory proteins from yeast, were faithfully translocated and glycosylated by Neurospora microsomes when presence cotranslationally in the Neurospora translation system. When presence cotranslationally in the Neurospora translation system, microsomes from canine pancreas(cRM) could also translocate and glycosylate the secretory proteins. However, salt-extracted cRM, which is depleted of canine signal recognition particle, could not. Furthermore, prepro alpha factor and a truncated form of invertase, containing the first 262-amino acid residues of the secretory invertase, were glycosylated by Neurospora microsomes posttranslationally, whereas only the truncated form of invertase was glycosylated by cRM when added posttranslationally. The full length invertase was not glycosylated posttranslationally. Posttranslational glycosylation of prepro alpha factor and of the truncated form of invertase is dependent on the hydrolysis of a nucleoside triphosphate. These data suggest that posttranslational glycosylation of prepro alpha factor occurs via a novel type of recognition mechanism which is either absent or ineffective in cRM.
D000818:Animals; D002473:Cell Wall; D004285:Dogs; D019286:Endopeptidase K; D005656:Fungal Proteins / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000378:metabolism; D006031:Glycosylation; D006596:Hexosaminidases / Q000378:metabolism; D000072235:Mating Factor; D008861:Microsomes / Q000378:metabolism; D008970:Molecular Weight; D009491:Neurospora / Q000378:metabolism*; D009492:Neurospora crassa / Q000235:genetics / Q000378:metabolism*; D010179:Pancreas / Q000378:metabolism / Q000648:ultrastructure; D010455:Peptides / Q000378:metabolism*; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D012265:Ribonucleotides / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000378:metabolism; D043324:beta-Fructofuranosidase
D005656:Fungal Proteins; D010455:Peptides; D012265:Ribonucleotides; D000072235:Mating Factor; D006026:Glycoside Hydrolases; D006596:Hexosaminidases; D043324:beta-Fructofuranosidase; D012697:Serine Endopeptidases; D019286:Endopeptidase K