A purification procedure is described yielding DNase I from bovine and rat parotid glands of high homogeneity. The apparent molecular masses of the DNases I isolated have been found by sodium dodecyl sulfate/polyacrylamide gel electrophoresis to be 34 and 32 kDa for bovine and rat parotid DNase I, respectively, and thus differ from the enzyme isolated from bovine pancreas (31 kDa). By a number of different criteria concerning their enzymic behaviour, the isolated enzymes could be clearly classified as DNases I, i.e. endonucleolytic activity preferentially on native double-stranded DNA yielding 5'-oligonucleotides, a pH optimum at about 8.0, the dependence of their enzymic activity on divalent metal ions, their inhibition by 2-nitro-5-thiocyanobenzoic acid and by skeletal muscle actin. Comparison of their primary structure by analysis of their amino acid composition and also two-dimensional fingerprints and isoelectric focusing indicate gross similarities between the enzymes isolated from bovine pancreas and parotid, but distinct species differences, i.e. between the enzymes isolated from bovine and rat parotid. All the DNases I are glycoproteins. From bovine parotid DNase I crystals suitable for X-ray structure analysis could be obtained. The DNases

Rat and bovine parotid gland and pancreas contain deoxyribonuclease I (DNAase I) activities in different amounts. The DNAase I activity in tissue homogenates of bovine and rat parotid gland can be inhibited by addition of monomeric actin, as with the enzyme of bovine pancreas. The isolated DNAase I species from bovine and rat parotid gland differ in their molecular weights and also in their affinities for monomeric actin, being lowest for rat parotid DNAase I (5 X 10(6)M(-1). Antibodies raised against rat and bovine parotid and bovine pancreatic DNAase I can be used to study the subcellular localization of DNAase I in these tissues by indirect immunofluorescence. DNAase I was found to be confined solely to the secretory granules of the tissue from which it was isolated.