A NOVEL HUMAN CHYMOTRYPSIN-LIKE DIGESTIVE ENZYME

The gene of a novel chymotrypsin-like serine protease has been cloned from human pancreas. The chymotrypsin-like enzyme-1 gene is located on chromosome 16q22.1 in a tight cluster with four unrelated genes. The gene has seven exons with the signal and activation peptide and the th ree main catalytic residues forming the active site encoded by separat e exons. Northern blots of pancreatic mRNA showed a major transcript o f 1.0 kilobases and a minor transcript of 1.3 kilobases due to alterna tive polyadenylation. No transcript was found in other tissues. Its pr esence in pancreatic tissue, duodenal juice, and urine was demonstrate d with antisera raised against synthetic peptides from the derived ami no acid sequence of the gene, The peptide sequences were chosen for be ing most dissimilar to chymotrypsin, and the antisera obtained did not react with purified human chymotrypsin. The proteolytically active CT RL-1 has been identified in pancreatic homogenate, duodenal juice, and urine, and a recombinant CTRL-1 has been characterized. Increased pan creatic secretion of CTRL-1 was induced by protease inhibitors indicat ing that the enzyme is secreted from pancreas upon feedback stimulatio n. Both native and recombinant CTRL-1 displayed chymotrypsin- and elas tase-2-like activities and hydrolyzed the amide bonds of substrates ha ving tyrosine, phenylalanine, or leucine residues at the p(1) position . The enzyme was active over a broad pH range (6.5-9.0), with a maximu m at pH 8.0-8.5. CTRL-1 was produced as a zymogen of 264 amino acids a s deduced from the gene sequence, with a sequence identity of 54% with human chymotrypsin B. The number and location of intron/exon junction s as well as the sequence similarity to chymotrypsin both at the DNA a nd protein level and the presence in duodenal juice indicate that this is a novel digestive enzyme of the chymotrypsin superfamily, albeit o ne with distinct physiological and biochemical features.