Hodel A, An S J, Hansen N J, Lawrence J, Wäsle B, Schrader M, Edwardson J M
Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QJ, UK.
Biochem J. 2001 Jun 15;356(Pt 3):843-50. doi: 10.1042/0264-6021:3560843.
Syncollin is a protein of the pancreatic zymogen granule that was isolated through its ability to bind to syntaxin. Despite this in vitro interaction, it is now clear that syncollin is present on the luminal side of the zymogen granule membrane. Here we show that there are two pools of syncollin within the zymogen granule: one free in the lumen and the other tightly associated with the granule membrane. When unheated or cross-linked samples of membrane-derived syncollin are analysed by SDS/PAGE, higher-order forms are seen in addition to the monomer, which has an apparent molecular mass of 16 kDa. Extraction of cholesterol from the granule membrane by treatment with methyl-beta-cyclodextrin causes the detachment of syncollin, and this effect is enhanced at a high salt concentration. Purified syncollin is able to bind to brain liposomes at pH 5.0, but not at pH 11.0, a condition that also causes its extraction from granule membranes. Syncollin binds only poorly to dioleoyl phosphatidylcholine liposomes, but binding is dramatically enhanced by the inclusion of cholesterol. Finally, cholesterol can be co-immunoprecipitated with syncollin. We conclude that syncollin is able to interact directly with membrane lipids, and to insert into the granule membrane in a cholesterol-dependent manner. Membrane-associated syncollin apparently exists as a homo-oligomer, possibly consisting of six subunits, and its association with the membrane may be stabilized by electrostatic interactions with either other proteins or phospholipids.
Syncollin是胰腺酶原颗粒中的一种蛋白质，它是通过其与 syntaxin 的结合能力而被分离出来的。尽管存在这种体外相互作用，但现在很清楚的是，syncollin 存在于酶原颗粒膜的腔面。在这里我们表明，在酶原颗粒中有两个syncollin池：一个游离在腔内，另一个与颗粒膜紧密结合。当通过SDS/PAGE分析未加热或交联的膜衍生syncollin样品时，除了表观分子量为16 kDa的单体之外，还可以看到高阶形式。用甲基-β-环糊精处理从颗粒膜中提取胆固醇会导致syncollin的脱离，并且在高盐浓度下这种作用会增强。纯化的syncollin能够在pH 5.0时与脑脂质体结合，但在pH 11.0时则不能，pH 11.0的条件也会导致其从颗粒膜中被提取出来。Syncollin与二油酰磷脂酰胆碱脂质体的结合很差，但通过加入胆固醇，结合会显著增强。最后，胆固醇可以与syncollin共免疫沉淀。我们得出结论，syncollin能够直接与膜脂质相互作用，并以胆固醇依赖的方式插入颗粒膜中。膜相关的syncollin显然以同型寡聚体的形式存在，可能由六个亚基组成，并且它与膜的结合可能通过与其他蛋白质或磷脂的静电相互作用而稳定。
