Analysis of the association of syncollin with the membrane of the pancreatic zymogen granule

Syncollin is a pancreatic zymogen granule protein that was isolated through its ability to bind to syntaxin, Here we show that syncollin has a cleavab le signal sequence and can be removed from granule membranes by washing wit h sodium carbonate. When membranes were subjected to Triton X-114 partition ing, syncollin was found predominantly in the aqueous phase, indicating tha t it is not sufficiently hydrophobic to be embedded in the membrane. Syncol lin has intramolecular disulfide bonds and was accessible to water-soluble cross-linking and biotinylating reagents only when granules were lysed by s onication. These results indicate that syncollin is tightly bound to the lu minal surface of the granule membrane. In situ, syncollin was resistant to proteases such as trypsin, When granule membranes were solubilized in ionic detergents such as deoxycholate, this trypsin resistance was maintained, a nd syncollin migrated on sucrose density gradients as a large (150 kDa) pro tein, In contrast, in non-ionic detergents such as Triton X-100, syncollin became partially sensitive to trypsin and behaved as a monomer. Syncollin i n alkaline extracts of granule membranes was also monomeric. However, reduc tion of the pH regenerated the oligomeric form, which was insoluble. We con clude that syncollin exists as a homo-oligomer and that its ability to self -associate can be reversibly modulated via changes in pH, In light of our f indings, we reassess the likely role of syncollin in the pancreatic acinar cell.