Structure of a Non-psychrophilic Trypsin from a Cold-Adapted Fish Species

Abstract

The crystal structure of cationic trypsin (CST) from the Atlantic salmon (Salmo salar) has been refined at 1.70 ,A, resolution. The crystals are orthorhombic, belong to space group P212121, with lattice parameters a = 65.91, b = 83.11 and c = 154.79 ,~,, and comprise four molecules per asymmetric unit. The structure was solved by molecular replacement with AMoRe and refined with X-PLOR to an R value of 17.4% and Rfree of 21.5% for reflections IFI > 30"F between 8.0 and 1.7 A resolution. The four non-crystallographic symmetry (NCS) related molecules in the asymmetric unit display r.m.s, deviations in the range 0.31-0.74,~ for main-chain atoms, with the largest differences confined to two loops. One of these is the calcium-binding loop where the electrondensity indicates a calcium ion for only one of the four molecules. In order to find structural rationalizations for the observed difference in thermostability and catalytic efficiency of CST, anionic salmon trypsin (AST) and bovine trypsin (BT), the three structures have been extensively compared. The largest deviations for the superimposed structures occur in the surface loops and particularly in the so-called 'autolysis loop'. Both the salmon enzymes possess a high methionine content, lower overall hydrophobicity and enhanced surface hydrophilicity, compared with BT. These properties have so far been correlated to cold-adaptation features, while in this work it is shown that the non-psychrophilic cationic salmon trypsin shares these features with the psychrophilic anionic salmon trypsin.

A Computational Module Assembled from Different Protease Family Motifs Identifies PI PLC from Bacillus cereus as a Putative Prolyl Peptidase with a Serine Protease Scaffold

“…PI-PLC was indicated to be a serine protease because the best match was with a trypsin protein, PDBid:1A0J [29]. The residues predicted by CLASP as responsible for its protease activity coincide with the active site responsible for its native phospholipase activity (His32, Asp67, His82, and Asp274) (Fig.…”

Trypsins from fish processing waste: characteristics and biotechnological applications – comprehensive review

“…Trypsin and trypsin-like proteolytic enzymes have been purified and characterized in several fish species including stomachless bone fish Carassius auratus gibelio (Bloch) (Jany, 1976), sardine (Murakami and Noda, 1981), capelin (Hjelmeland and Raa, 1982), Greenland cod (Simpson et al, 1989), cunner (Simpson et al, 1989), Atlantic cod (Amiza et al, 1997;Asgeirsson et al, 1989;Han, 1993;Simpson et al, 1989), chum salmon (Uchida et al, 1984a(Uchida et al, , 1984b, Atlantic salmon (Male et al, 1995;Schroder et al, 1998), coho salmon (Haard et al, 1996), anchovy (Martinez et al, 1988), Atlantic white croaker (Pavlisko et al, 1997b), carp (Cyprinus carpio) (Cao et al, 2000), arabesque greenling (Pleuroprammus azonus) (Table 1).…”

Structural Analysis of the Human Golgi-associated Plant Pathogenesis Related Protein GAPR-1 Implicates Dimerization as a Regulatory Mechanism

“…C, The two molecules of the proposed dimer share a continuous hydrophobic core (only Trp and Tyr shown). D, The position of the conserved amino acid residues proposed to form a catalytic triad between the two molecules at the end of a groove running through the centre of the dimer (superimposed catalytic residues from trypsin are shown coloured in red; PDB: 1A0J 48 ). E, A close up of the catalytic triad compared with those from trypsin (superimposed catalytic residues from trypsin are shown coloured in red; PDB: 1A0J 48 ).…”