A new substrate for enteropeptidase

Abstract

Enteropeptidase (EC 3.4.4.8) a glycoprotein of mol. wt 200 000 is an enzyme of the small intestine [l-5] . Its physiological function is to catalyse the formation of trypsin from pancreatic trypsinogen, by the removal of a small amino-terminal peptide (trypsinogen activation peptide). The trypsin formed in this manner is essential for the activation of other pancreatic zymogens and the digestion of food proteins by cooperative enzymatic action [6]. Current assay methods for enteropeptidase activity [4,7-91 are based on the activation of trypsinogen and the subsequent splitting of an artificial substrate by trypsin. These methods are sensitive, but inherently complex, careful control of the assay conditions being necessary in order to avoid autoactivation and autodegradation of trypsinogen. In addition, the proteolytic action of trypsin upon the assayed material must be regarded as a serious drawback in subcellular localisation studies. An assay method for enteropeptidase activity based on the hydrolysis of a specific artificial substrate would therefore have the double advantage of simplifying the assay procedure and removing uncertainties due to the generation of trypsin in the incubation mixture. The amino acid sequence of activation peptides of trypsinogen from different species has been reported [lo] . The partial sequence, -Asp-Asp-Asp-Asp-Lys has been shown by Maroux et al. [7] to comprise the