Cholesterol-dependent interaction of syncollin with the membrane of the pancreatic zymogen granule.
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.
Hodel
A
A
Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QJ, UK.
An
S J
SJ
Hansen
N J
NJ
Lawrence
J
J
Wäsle
B
B
Schrader
M
M
Edwardson
J M
JM
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002784:Cholesterol / Q000378:metabolism*; D003594:Cytoplasmic Granules / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D010179:Pancreas / Q000378:metabolism*; D011233:Precipitin Tests; D051381:Rats
D002352:Carrier Proteins; D008565:Membrane Proteins; C107454:Sycn protein, rat; D002784:Cholesterol