On the preparation and some properties of closed membrane vesicles from hog duodenal and jejunal brush border

Abstract

Closed and nearly spherical vesicles were obtained from both hog duodenum and jejunum after mucosa homogenization in the absence of EDTA and a series of fractional centrifugations. The vesicles were found to contain large amounts of two of the characteristic enzyme markers of the brush border membrane (aminopeptidase and alkaline phosphatase). They were seen by electron microscopy on thin sections or after negative staining to be composed of an apparently intact, 90–100 Å-thick membrane overlaid by the fuzzy coat and to be partly filled by a fibrous material tentatively identified with the cross-filaments of the microvilli. This filling was not removed by 5 mM EDTA or/and 1 M Tris unless the structure of the vesicles was largely destroyed. Very few empty vesicles were obtained at the end of these treatments. The vesicles from hog duodenum and jejunum were observed to contain nearly 2 molecules of cholesterol for 1 molecule of phospolipids. Specific differences were noted between both types of vesicles at the level of their sugar composition and associated enzyme activities. For instance, the jejunal vesicles contained no sialic acid and no enterokinase. They contain, respectively, 2 and 4 times as much alkaline phosphatase and aminopeptidase as duodenal vesicles.