A NEW SITE-SPECIFIC ENDONUCLEASE METHYLASE FROM THE THERMOPHILIC STRAIN BACILLUS SPECIES LU11

A new site-specific endonuclease, BspLU11III, was isolated and purified to homogeneity from the thermophilic strain Bacillus species LU11. The enzyme recognizes the 5'-GGGAC-3' sequence on double-stranded DNA and cleaves it at two places, 10 and 11 nucleotides from the 3'-end of the 5'-GGGAC-3' sequence and 14 and 15 nucleotides from the recognized site along the complementary strand. In solution the enzyme is a monomer with molecular mass of about 93 kD. In the presence of S-adenosylmethionine the enzyme has methylase activity. The adenine residue in the recognition site is methylated on one of the DNA strands. The restriction activity of BspLU11III does not depend on ATP, but is stimulated by 80 mu M S-adenosylmethionine. Mg2+ is required for restriction activity. Star activity is observed in the presence of sodium chloride. BspLU11III is not a member of the three main classes of endonucleases, but has properties similar to type IV site-specific endonucleases.