Endonuclease I of Proteus mirabilis: II. PROPERTIES OF THE NONCOMPLEXED AND TRANSFER RIBONUCLEIC ACID-COMPLEXED FORMS OF THE ENZYME

Abstract

Abstract The purified form of endonuclease I of Proteus mirabilis sediments as a 3 S molecule, requires Mg2+ for activity, and degrades double and single stranded DNA to acid-soluble products. This activity has an alkaline pH optimum and is inhibited by Escherichia coli tRNA and high concentrations of sodium chloride. The two faster sedimenting forms of the enzyme, 7.2 and 6 S, that can be generated by the addition of tRNA to the purified enzyme and also are found in a dihydrostreptomycin sulfate precipitate of a P. mirabilis crude extract, degrade the covalently closed circular duplex form of DNA of colicinogenic factor E1, φ X-174, and SV40 DNA to 8 S DNA fragments that are indistinguishable by sucrose gradient centrifugation analysis. Degradation of the supercoiled circular DNA to the 8 S product is independent of substrate concentration over a wide range and is not caused by inactivation of the enzyme under the reaction conditions used. The 8 S DNA product is double stranded, free of a substantial amount of nicks, and, once formed, is resistant to further endonuclease attack over a relatively long period of time under the conditions used. Evidence is presented for a single strand cleavage as the initial endonucleolytic event in the degradation of the native DNA substrate to the 8 S product.