title: Calnexin and calreticulin bind to enzymically active tissue-type plasminogen activator during biosynthesis and are not required for folding to the native conformation.
abstract: The roles of the endoplasmic-reticulum lectins calnexin and calreticulin in the folding of tissue-type plasminogen activator (tPA) have been investigated using an in vitro translation system that reconstitutes these processes as they would occur in the intact cell. Using co-immunoprecipitation of newly synthesized tPA with antibodies to calnexin and calreticulin, it was demonstrated that the interaction of tPA with both lectins was dependent upon tPA glycosylation and glucosidase trimming. When tPA was synthesized in the presence of semi-permeabilized cells under conditions preventing complex formation with calnexin and calreticulin, the translation product had a specific plasminogenolytic activity identical with that when synthesized under conditions permitting interactions with both lectins. Furthermore, complexes of tPA bound to calnexin and calreticulin were shown to be enzymically active. These results demonstrate that calnexin and calreticulin can form a stable interaction with correctly folded tPA; however, such interactions are not required for the synthesis of enzymically active tPA.
authors.affiliation: University of Manchester, School of Biological Sciences, UK.
authors.forename: S
authors.initials: S
authors.lastname: Allen
authors.forename: N J
authors.initials: NJ
authors.lastname: Bulleid
mesh_terms: D002135:Calcium-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D037281:Calnexin; D037282:Calreticulin; D002460:Cell Line; D002463:Cell Membrane Permeability; D006031:Glycosylation; D006801:Humans; D046911:Macromolecular Substances; D009844:Oligosaccharides / Q000378:metabolism; D010958:Plasminogen / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D012261:Ribonucleoproteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D013601:T-Lymphocytes / Q000201:enzymology / Q000378:metabolism; D010959:Tissue Plasminogen Activator / Q000737:chemistry / Q000378:metabolism*
chemical_list: D002135:Calcium-Binding Proteins; D037282:Calreticulin; D046911:Macromolecular Substances; D009844:Oligosaccharides; D012261:Ribonucleoproteins; D037281:Calnexin; D010958:Plasminogen; D010959:Tissue Plasminogen Activator