A site-specific endonuclease from the thermophilic strain Bacillus speciesF4

A strain producing the site-specific endonuclease BspF4I was found during s creening of thermophilic bacteria isolated from soil. The restriction endon uclease, free from contaminant nonspecific nucleases, was purified using th ree steps of column chromatography-on hydroxyapatite, blue agarose, and DEA E-Trisacryl. The enzyme is stable on storage and exhibits maximal activity at 48-56 degrees C in the presence of albumin in buffer containing 10 mM Tr is-HCl (pH 7.5) and 10 mM MgCl2. BspF4I recognizes the sequence 5'GGNCC-3' on DNA and is an isomer and not an isoschizomer of the endonuclease Sau96I. Unlike the prototype, BspF4I does not cleave the site in a defined way. A strand with purine in the center of the sequence is cleaved after the first G, as in the case of the prototype, while the strand with pyrimidine is cl eaved either before or after the first G.