A DNA endonuclease isolated from yeast nuclear extract

Abstract

We have isolated and partially purified a DNA endonuclease from nuclei of the yeast Saccharomyces cerevisiae. Although purified on the basis of its ability to degrade denatured DNA, the enzyme can also attack native DNA. Denatured oligonucleotide products of the enzyme are sensitive to venom phosphodiesterase (EC 3.1.4.1) but not to bovine spleen phosphodiesterase (EC 3.1.4.18). The enzyme has an estimated molecular weight of 6.6–7.5 × 104, more than twice as large as the endonucleases involved in DNA repair in Escherichia coli.When analyzed on glycerol gradients, the endonuclease sedimented as a single activity against both denatured DNA and closed circular DNA duplexes. The enzyme showed a 10-fold preference for denatured over native T7 DNA substrate, and appears to produce random nicks in a supercoiled replicative form of [Formula: see text] DNA (RFI) with no discernable preference for the unpaired bases in the supercoiled duplex. The endonuclease appears to be distinct from the yeast endonucleases previously described.