Residue determinants and sequence analysis of cold-adapted trypsins

The digestive enzyme trypsin is among the most extensively studied proteins , and its structure has been reported from a large number of organisms. Thi s article focuses on the trypsins from vertebrates adapted to life at low t emperatures. Cold-adapted organisms seem to have compensated for the reduce d reaction rates at low temperatures by evolving more active and less tempe rature-stable enzymes. We have analyzed 27 trypsin sequences from a variety of organisms to find unique attributes for the cold-adapted trypsins, comp aring trypsins from salmon, Antarctic fish, cod, and pufferfish to other ve rtebrate trypsins. Both the "cold" and the "warm" active trypsins have abou t 50 amino acids that are unique and conserved within each class. The main unique features of the cold-adapted trypsins attributable to low-temperatur e adaptation seem to be (1) reduced hydrophobicity and packing density of t he core, mainly because of a lower (Ile + Leu)/(Ile + Leu + Val) ratio, (2) reduced stability of the C-terminal, (3) lack of one warm trypsin conserve d proline residue and one proline tyrosine stacking, (4) difference in char ge and flexibility of loops extending the binding pocket, and (5) different conformation of the "autolysis" loop that is likely to be involved in subs trate binding.