Dog pancreatic microsomal-membrane polypeptides analysed by two-dimensional gel electrophoresis.
The two-dimensional polyacrylamide-gel electrophoresis technique of O'Farrell [(1975) J. Biol. Chem 250, 4007-4021] was applied to resolve and analyse the polypeptide composition of dog pancreatic rough microsomal membranes, which were shown to be active in co-translational processing of preprolactin synthesized from pituitary mRNA in a translation system in vitro. About 100 polypeptides are resolved. Treatment of rough microsomal membranes with EDTA and high KCl concentration yielded membranes stripped of their ribosomes with retention of activity for translocation and processing. Stripped microsomal membranes showed a selective concentration of approximately 25 polypeptides in the membranes when analysed by two-dimensional polyacrylamide-gel electrophoresis. The two-dimensional electrophoretic profile was catalogued into polypeptides that are glycoproteins, those that contain free thiol groups disposed at the cytosolic surface of microsomal vesicles and those that are of secretory origin but have been entrapped in the microsomal preparation. Several secretory components, including amylase, procarboxypeptidases, lipase and anionic trypsinogen, were tentatively identified among the microsomal polypeptides. The rough and stripped microsomal membranes from dog pancreas show a characteristic set of seven major acidic polypeptides, which are also identifiable in microsomal-membrane preparations isolated from dog liver and rat liver. One of these polypeptides was identified as protein disulphide-isomerase (EC 5.3.4.1).
D000818:Animals; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D007425:Intracellular Membranes / Q000032:analysis; D007525:Isoelectric Focusing; D007535:Isomerases / Q000032:analysis; D008565:Membrane Proteins / Q000032:analysis; D008861:Microsomes / Q000032:analysis; D008862:Microsomes, Liver / Q000032:analysis; D010179:Pancreas / Q000032:analysis*; D010455:Peptides / Q000032:analysis*; D019704:Protein Disulfide-Isomerases; D051381:Rats; D013050:Spectrometry, Fluorescence
D008565:Membrane Proteins; D010455:Peptides; D007535:Isomerases; D019704:Protein Disulfide-Isomerases