The third serine proteinase with chymotrypsin specificity isolated from Atlantic cod (Gadus morhua) is a type‐II elastase

Abstract

Preparations of chymotrypsin from Atlantic cod are heterogeneous and previously gave rise to two active peaks when subjected to pH-gradient chromatography. Extension of the pH-gradient resolved a third protein peak with benzoyltyrosine ethylester hydrolytic activity. The first two peaks have been characterized as chymotrypsin variants and designated A and B, whereas the identity of the third peak was not clear. Analysis of this protein by Edman sequencing and mass spectrometry has now confirmed a high degree of identity with the predicted protein sequence from a recently described cDNA clone. That sequence was named elastase B by sequence comparison. As the present elastase deviates in 16 positions from that of elastase B, we have named it elastase C. The elastase C was active in hydrolysing typical substrates used by chymotrypsin, namely benzoyl-L-tyrosine ethylester and succinyl-Ala-Ala-ProPhe-p-nitroanilide, but inactive against the typical elastase substrates succinyl-Ala-Ala-Ala-p-nitroanilide and orcein-elastin. Comparison of the kinetic properties of the cod elastase C with bovine chymotrypsin and cod chymotrypsin variants A and B, using succinyl-Ala-Ala-Pro-Phe-p-nitroanilide, showed a lower catalytic efficiency of elastase C. The effects of several inhibitors on cod elastase C were identical to effects on chymotrypsins variants A and B, but dissimilar when compared with porcine pancreatic elastase. On the basis of the specificity and amino acid sequence, we conclude that the enzyme under study is most correctly classified as a type-II elastase.Keywords : elastase; serine protease ; Atlantic cod ; amino acid sequence; cold adaptation.Variants of elastases have been identified in different animals and in different animal tissues such as the pancreas, leucocytes, platelets, and aorta [1]. Elastase was initially characterized as a mammalian pancreatic serine protease with the ability to dissolve elastin [2], but elastase can digest a wide variety of other protein substrates in addition to native elastin [3]. Some elastases in the serine proteinases family turn out to be specific for non-bulky amino acid residues (e.g. porcine pancreatic elastase 1, human leucocyte elastase) while others have a chymotrypsinlike specificity (e.g. porcine pancreatic elastase 2, human pancreatic elastase 2). Pancreatic elastase type I was originally isolated from hog pancreas and is strongly cationic with an isoelectric point near pH 9.5 [2]. Another anionic enzyme, called elastase type II, was also obtained from the same tissue and found to have a characteristic chymotrypsin activity [4,5]. Human pancreatic tissue and juice also contain two enzymes, one cationic and the other anionic, which can hydrolyse a specific substrate for porcine pancreatic elastase and undyed elastin [6,7]. The cationic enzyme is the major elastase in human pancreas and has similar properties to porcine pancreatic elastase 1, whereas elastase 2 preferentially hydrolysed peptides at sites of medium to large hydrophobic side chains [8]. Human elastase 1 ...

Characterization of proteases in guts ofDaphnia magnaand their inhibition byMicrocystis aeruginosaPCC 7806

“…When the chymostatin-specific substrate S(Ala) 2 Pro-PhepNA was used, only chymostatin inhibited the proteolysis by D. magna gut homogenate. Another difference found was that elastatinal, which is a specific inhibitor of mammalian elastase, inhibited hydrolysis of the chymotrypsin substrate S(Ala) 2 ProPhepNA in D. magna gut homogenates; a similar finding of a chymotrypsin with characteristics of elastase was recently reported for Atlantic cod (Asgeirsson et al, 1998). Hence, D. magna contains digestive chymotrypsins that differ from vertebrate chymotrypsins in both their sensitivity to inhibitors and their specificity for substrates.…”

Utilization of marine by-products for the recovery of value-added products

“…However, Shotton (1970) reported that elastase can digest a wide variety of other proteins in addition to native elastin. Gastric elastase enzymes have been recovered from marine and fresh water fish species such as carp (Cohen and Gertler, 1981), catfish (Clark, et al, 1985), and Atlantic cod (Asgeirsson and Bjarnason, 1993;Asgeirsson, et al, 1998;Gildberg and Overbo, 1990;Raa and Walther, 1989). Apart from the collagenases distributed in digestive tract, they were also found in fish muscle tissues including skeletal muscle of mackerel, Scomber japonicus, Japanese flounder, Paralichthys olivaceus, rainbow trout and common carp (Sriket, 2014).…”