New Site-Specific Endonucleases F-TflI, F-TflII, and F-TflIV Encoded by Bacteriophage T5

Site-specific endonucleases F-TflI, F-TflII, and F-TflIV have been revealed, which belong to the H-N-H family and are encoded by ORFs located in the tRNA gene region of bacteriophage T5. It has been shown that endonuclease F-TflIV introduces a double-strand break in a 17-bp pseudopalindromic DNA sequence to yield 1-nt 3′-protruding ends. Unlike F-TflIV, F-TflI, and F-TflII introduce single-strand breaks in asymmetrical, highly degenerate sequences, each cleaving only one (template or coding) strand. Amino acid sequence analysis has revealed a high homology of the enzymes in the region of the H-N-H motif and in the putative C-terminal catalytic domain. The N-terminal region of F-TflIV proved to be homologous to the HTH domain of LuxR-related transcriptional regulators, which is responsible for DNA recognition and binding. The N-terminal regions of F-TflI and F-TflII contain a composite motif NUMOD4, which is characteristic of a putative recognition domain of some H-N-H endonucleases. A two-domain structure, with the N-terminal recognition and C-terminal catalytic domains, and evolutionary origin via recombination of the catalytic and recognition domain-coding regions are proposed for F-TflI, F-TflII, and F-TflIV.