Protein digestion in human and rat small intestine: Role of new neutral endopeptidases

Abstract

Two new phosphoramidon-insensitive neutral endopeptidases were identified and partially characterized in the brush-border membrane of rat and human intestine using N-CBZ-L-Ala-L-Arg-L-Arg-4-methoxy-β-naphthylamide (Z-Ala-Arg-Arg-MNA) and azocasein or α-casein as substrates. Activities in the brush-border membrane of both rat and human intestine were maximum at neutral to alkaline pH, were inhibited by metal chelating and thiol reagents, and were insensitive to phosphoramidon. The results also indicate that these endopeptidases are distinct from pancreatic proteases. The biochemical properties of the enzyme hydrolyzing Z-Ala-Arg-Arg-MNA were shown to be different from that hydrolyzing azocasein or α-casein. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration revealed that several native intact protein substrates were rapidly degraded to small molecular weight peptides and amino acids when incubated with rat or human brush-border membrane preparations. During in vivo intestinal perfusion in rats, 11% of the total administered α-casein was hydrolyzed and absorbed by the intestine. The results suggest that phosphoramidon-insensitive endopeptidases in the intestinal brush-border membrane may be of nutritional and physiological importance in protein digestion.