The effects of variation in dietary protein content have been investigated on brush border glycosylation and enzyme activities in mice small intestine. The comparison of different parameters was made between the mice fed 30% (high protein, HP) and 18 % protein (pair-fed, PF, and ad libitum-fed) for 21 days. The activities of brush border sucrase, lactase, p-nitrophenyl (PNP)-β-D-glucosidase and PNP-β-D-galactosidase were reduced in the HP diet-fed mice compared to PF and ad libitum-fed controls. Alkaline phosphatase and leucine aminopeptidase activities were significantly enhanced while γ-glutamyl transpeptidase activity was unaltered under these conditions. Total hexoses and sialic acid content in the brush borders were reduced significantly in the test group compared to the controls while hexosamine and fucose contents remained essentially similar in different groups. The results on the binding of wheat germ agglutinin and Ulex europaeus agglutininI to microvillus membranes corroborated the chemical analysis data on sialic acid and fucose contents of the membranes. Peanut agglutinin binding was enhanced in mice from the HP group. Incorporation of (14C)-mannose into membranes was significantly less in HP diet-fed mice. These results indicate that the feeding of HP diet to mice brings about marked alterations in small intestinal epithelial cell surface glycosylation and enzyme functions.