Characterization of a pancreatic DNase from pyloric caeca of atlantic cod (Gadus morhua L.).

An alkaline deoxyribonuclease (DNase) from cod pancreatic tissue has been characterized. The enzyme is a DNase I type endonuclease and hydrolyzes effectively both native and denatured DNA. Monomeric actin inhibits the enzyme reaction. The enzyme obeys Michaelis-Menten kinetics and the apparent Km value for native linear duplex DNA is 33 µg/ml. The cod DNase opens supercoiled plasmid DNA, by introducing adjacent nicks in both strands, possibly separated by 5-10 nucleotides. DNA hydrolyzed by cod DNase functions as substrates both for DNA polymerase and ligase, and the nicks therefore contain 5'-phosphoryl and 3'-hydroxyl groups. Optimum concentrations of divalent cations are 5 mM Mg(2+), 0.63 mM Mn(2+) and 0.075 mM Ca(2+). However, Ca(2+) is apparently not essential for the enzymatic functions. The enzyme has a narrow temperature optimum at 42°C and is thermolabile above 50°C; however, Mn(2+) shifts the optimum slightly to 45°C by causing increased temperature stability. The cod DNase reaction is inhibited by the DNA intercalating compounds actinomycin D and ethidium bromide. Histidine-modifying reagents such as tosyl phenylalanyl chloromethylketone and diethyl pyrocarbonate inhibit the enzyme activity, but the cod DNase is insensitive to disulfide-reducing agents.