Partial purification and properties of a nucleoside triphosphate-dependent deoxyribonuclease from Mycobacterium smegmatis

Abstract

An ATP-dependent deoxyribonuclease was partially purified from iron-limited Mycobacterium smegmatis. The enzyme showed a strong preference for native DNA, and was completely dependent on the presence of a riboside, or deoxyriboside, triphosphate and Mg 2+. Maximal activity was reached with about 1 mg of sodium DNA per ml, 4 mM ATP and 30 mM Mg 2+. In the presence of DNA the enzyme hydrolyzed ATP to ADP and orthophosphate, one mole of ATP being hydrolyzed per mole of deoxyribonucleotide solubilized. The DNA was attacked from the ends of strands, and the evidence favoured attack on both strands. The products released from DNA were 4–13 % mononucleotides, 14–26 % dinucleotides, 29–45 % trinucleotides, 15–23 % tetranucleotides and 5–29 % of longer oligonucleotides, as fractionated on DEAE-Sephadex. Fractionation on Sephadex G-25, on DEAE-paper, and by two-dimensional paper chromatography gave results in keeping with this. The properties of the enzyme are discussed in the light of a possible role in recombination.