Rat intestinal microvillus membranes. Purification and biochemical characterization

Abstract

A technique is described for the removal of subcellular contaminants from intact rat intestinal brush borders, and for the subsequent separation of a microvillus membrane fraction from a fibrillar residue. Increments in invertase activity, microscopic homogeneity and low nucleic acid content indicated that the microvillus plasma membrane was extensively purified. Multiple membrane preparations were highly reproducible with respect to their invertase specific activity, cholesterol content and phospholipid content. Alkaline phosphatase, leucine aminopeptidase, Mg2+- and Ca2+-dependent ATP and 7 separate disaccharidases were predominantly confined to the membrane fraction. The fibrillar fraction contained approximately 30% of the total protein of purified brush borders, plus most of the residual nucleic acid contaminant. No evidence was found for the localization of any specific enzyme in this fraction.