Enterokinase, the initiator of intestinal digestion, is a mosaic protease composed of a distinctive assortment of domains.

Abstract

Enterokinase is a protease of the intestinal brush border that specifically cleaves the acidic propeptide from trypsinogen to yield active trypsin. This ITPK-IVGG (human) or VSPK-IVGG (bovine), suggesting that single-chain enterokinase is activated by an unidentified trypsin-like protease that cleaves the indicated Lys-fle bond. Therefore, enterokinase may not be the "first" enzyme of the intestnal digestive hydrolase cascade. The specificity of enterokinase for the DDDDK-I sequence of trpsinogen may be explained by complementary basic-amino acid residues clustered in potential S2-S5 subsites.All animals need to digest exogenous macromolecules without destroying similar endogenous constituents. The regulation of digestive enzymes is, therefore, a fundamental requirement (1). Vertebrates have solved this problem, in part, by using a two-step enzymatic cascade to convert pancreatic zymogens to active enzymes in the lumen of the gut. The basic features ofthis cascade were described in 1899 by N. P. Schepovalnikov, working in the laboratory of I. P. Pavlov (2). Extracts of the proximal small intestine were shown to strikingly activate the latent hydrolytic enzymes in pancreatic fluid. Pavlov considered this intestinal factor to be an enzyme that activated other enzymes, or a "ferment of ferments," and named it "enterokinase." The importance of this protease cascade is emphasized by the life-threatening intestinal malabsorption that accompanies congenital deficiency of enterokinase (3, 4).Enterokinase activates bovine trypsinogen by cleaving after the sequence VDDDDK, releasing an amino-terminal activation peptide (5,6). The acidic DDDDK sequence of the trypsinogen-activation peptide is conserved among vertebrates (7), except for the similar sequences of trypsinogens from lungfish (IEEDK and LEDDK) and African clawed frog (FDDDK). Enterokinase prefers substrates with the sequence DDDDK, whereas the presence of aspartate residues markedly inhibits the ability of trypsin to cleave such substrates (8). For example, toward bovine trypsinogen the catalytic efficiency of enterokinase is 12,000-fold (porcine) (9) or 34,000-fold (bovine) (10) greater than that of bovine trypsin. This reciprocal specificity protects trypsinogen against autoactivation by trypsin and promotes activation by enterokinase in the gut.Enterokinase has been purified from porcine (11), bovine (10, 12, 13), human (14), and ostrich intestine (15). With the possible exception of human enterokinase, which was suggested to be a heterotrimer (14), enterokinase appears to be a disulfide-linked heterodimer with a heavy chain of 82-140 kDa and a light chain of 35-62 kDa. Mammalian enterokinases contain 30-50%o carbohydrate, which may contribute to the apparent differences in polypeptide masses. The heavy chain is postulated to mediate association with the intestinal brush border membrane (16), although no direct evidence for this function has been reported. The light chain contains the catalytic center. Based on susceptibility to inhibition by chem...

The astacin family of metalloendopeptidases

“…The MAM domain is found in several membrane proteins and has been proposed to act as an adhesion domain . Enteropeptidase (also termed enterokinase; EC 3.4.21.9), the intestinal enzyme that converts trypsinogen to trypsin, also contains a MAM domain (Kitamoto et al, 1994). MAM domains are 120-180 amino acids in length, and there are 14 amino acids, including 4 cysteine residues, that are conserved.…”

Molecular Cloning of cDNA for Matriptase, a Matrix-degrading Serine Protease with Trypsin-like Activity

“…These segments resemble partial sequences, originally identified in complement subcomponents C1r (25,26) and C1s (27,28). This C1r/s domain was also found in other serine proteases, such as enteropeptidase, an activator of trypsinogen (15,16), and in the astacin subfamily of zinc metalloprotease, such as bone morphogenetic protein-1 (29) and Drosophila tolloid gene, a dorsal-ventral patterning protein (30). Although the exact roles of the C1r/s domains in these proteins remain unclear, a deletion of the first C1r/s domain in complement subcomponent C1r impairs tetramer formation of C1r with C1s (31).…”

“…By analogy, the four tandem cysteine-containing repeats in matriptase may also be the sites of interaction with other macromolecules. In addition, the cysteine-containing LDL receptor domain was found in other proteases such as enteropeptidase (15,16).…”

“…The most likely cysteine pairings in matriptase are thus as follows: Cys-469 -Cys-485, Cys-604 -Cys-618, and Cys-629 -Cys-658). Matriptase also contains two additional cysteines (Cys-432-Cys-559) that correspond to those used in two-chain proteases, such as enteropeptidase (15,16), hepsin (17), plasma kallikrein (18), blood coagulation factor XI (19), and plasminogen (20), but not in trypsin (21) or chymotrypsin (22) (Fig. 4).…”

Prosegment of Tripeptidyl Peptidase I Is a Potent, Slow-binding Inhibitor of Its Cognate Enzyme

“…only two aa residues are found in granzymes (13), and as many as several hundred residues are found in enterokinase (14). The length of TPP I prosegment (176 aa) suggests that it could easily fold into an independent domain, and thus, in addition to active site shielding, it might also have other functions (i.e.…”