Comparison of the Multiple Forms of Bovine Pancreatic Deoxyribonuclease

Abstract The earlier resolution of preparations of bovine pancreatic deoxyribonuclease into two active fractions by chromatography on sulfoethyl-Sephadex has been improved through the use of columns of phosphocellulose. Gradient elution with sodium acetate at pH 4.7 shows the presence of at least four forms of the enzyme. The principal component (DNase A) is accompanied by Fractions B and C, and, in very minor amount, D; all four fractions possess comparable specific activities. Pancreatic juice contains DNases which are chromatographically indistinguishable from the A, B, and C components extracted from the tissue. The three main fractions (A, B, and C present in the extracted enzyme in the molar proportions 4:1:1) were analyzed for amino acids, terminal residues, and carbohydrates. All three proteins have NH2-terminal leucine, COOH-terminal threonine, and carbohydrate attached at a single position. DNases A and B have the same amino acid composition, but the B fraction differs from A (which possesses 2 N-acetylglucosamine and 6 mannose residues) in that it contains predominantly a DNase with 1 residue each of sialic acid and galactose in addition to 3 N-acetylglucosamine and 5 mannose residues. DNase C is similar to A in possessing a neutral carbohydrate moiety (2 N-acetylglucosamine and 5 mannose) but the protein contains 1 less histidine residue and 1 more proline residue than DNase A or B. Pancreatic tissue thus synthesizes DNases which vary in the carbohydrate side chain and in amino acid sequence.