Chymotrypsins from the deer (Cervidae) family. Isolation, partial characterization and primary-structure studies of chymotrypsins A and B from both moose (Alces alces) and elk (Cervus elaphus) pancreas.

• Main Authors: Lindsay, R M, Stevenson, K J
• Format: Text
• Language: English
• Published: 1976
• Subjects: Research Article; Alces alces
• Online Access: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1172877; http://www.ncbi.nlm.nih.gov/pubmed/949318

1. An anionic and a cationic chymotrypsin (EC 3.4.21.1) were isolated from the pancreas glands of the moose (Alces alces) and elk (Cervus elaphus). The A and B chymotrypsins from each species were purified to homogeneity by (NH4)2SO4 fractionation, affinity chromatography on 4-phenylbutylamine-Sepharose and ion-exchange chromatography on DEAE- and CM-cellulose. 2. The molecular weight and pH optimum of each chymotrypsin were similar to those of the corresponding ox A and B chymotrypsins. 3. The substrate specificities of the chymotrypsins were investigated by digestion of glucagon and the oxidized B chain of insulin. The primary specificity of each chymotrypsin for aromatic amino acid residues was further established by determining the Km and kcat for the hydrolysis of a number of synthetic amino acid ester substrates. 4. The amino acid composition and total number of residues of moose and elk chymotrypsin A were similar to those of ox chymotrypsin A. An even greater similarity was observed among the B chymotrypsins of the three species. 5. The A chymotrypsins of moose and elk were fragmented to their constituent 'A', 'B' and 'C' polypeptide chains by succinylation (3-carboxypropionylation), reduction and alkylation of the native enzymes. In each case, the two major chains ('B' and 'C') were separated and isolated. By comparison of the amino acid compositions of moose, elk and oxy 'B' and 'C' chains, a greater difference was observed among the three A chymotrypsins than was suggested by the amino acid compositions of the native enzymes alone. 6. Peptides were isolated from the disulphide bridge and active-site regions of the A and B chymotrypsins of moose and elk by diagonal peptide-'mapping' techniques. From the amino acid compositions of the isolated peptides (assuming maximum homology) and from a comparison of diagonal peptide 'maps', there was established a high degree of primary-structure identity among the mooae, elk and ox chymotrypsins. Tentative sequences were deduced for the peptides isolated by ...