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Cloning and characterization of a new site specific methyl- directed DNA endonuclease EcoBLI recognizing 5’- G(5mC)NGC-3’/3’-CGN(5mC)G-5’

D.A.Gonchar, V.A.Chernukhin, M.A.Abdurashitov, E.V.Kileva, V.S.Dedkov, N.A.Mikhnenkova, E.N.Lomakovskaya, S.G.Udalyeva, S.Kh. Degtyarev 
BioTechnology : An Indian JournalVolume 12,Issue 4, Pages 175-181

A gene coding BisI, site specific 5mC-directed DNA endonuclease recognizing DNA sequence 5’-G(5mC)NGC- 3’/3’-CGN(5mC)G-5’, was recently identified in the sequenced genome of the strain-producer Bacillus subtilis T30. In this work we have undertaken a search of bisI gene homologues among the sequenced genomes of enterobacteria. DNA analysis has revealed a small group of highly homologous ORFs with unknown function including one ORF in DNA of well-known strain E.coli. This ORF WP 001276099.1 from E.coli BL21 (DE3) was amplified and cloned. An obtained recombinant strain E.coli pEcoBLI produces MD-endonuclease named EcoBLI. The new enzyme has the same substrate specificity as BisI MD-endonuclease. Thus, ORF WP 001276099.1 from E.coli BL21 (DE3) encodes site-specific 5mC-directed DNA-endonuclease EcoBLI recognizing and cleaving DNA sequence as indicated by arrows 5’-G(5mC)^NGC-3’/3’-CGN^(5mC)G-5’.

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