DNA Sequence restricted by Restriction Endonuclease AP from Haemophilus aphirophilus

Abstract

MANY bacterial strains contain strain-specific restriction endonucleases which degrade foreign DNA at a limited number of sites (refs 1–6; P. A. Sparp, B. Sugden and J. Sambrook, personal communication). The site-specific action of these enzymes is thought to be a consequence of their ability to recognise specific nucleotide sequences⁵. Such sequences restricted by endonuclease R (endo R) from Haemophilus influenzae Rd (ref. 8) and RI endonuclease (endo RI) from Escherichia coli carrying R factor⁶ have been determined. In our laboratory, several species of similar enzymes have been isolated from different Haemophilus strains^{9, 10} Different enzymes seem to have different cleavage site specificities, since each enzyme produces different sizes of fragments from bacteriophage DNA¹⁰. We analysed the terminal nucleotide sequences of fragments produced from fd RF-I (doubly closed replicative form) DNA and T3 DNA by cleavage with one of the Haemophilus enzymes, endonuclease AP (endo AP) isolated from H. aphirophilus, and found that this enzyme cleaved DNA at a sequence of four nucleotide pairs with a two-fold rotational symmetry.