Strand specificity of nicking of DNA at Chi sites by RecBCD enzyme. Modulation by XXXX and magnesium levels.

@entity855 @entity1025 @entity526 @entity2036
RecBCD enzyme is essential for the major pathway of homologous recombination of linear DNA in @entity526 . It is a potent nuclease and helicase and, during its unwinding of double-stranded DNA, makes single-strand scissions in the vicinity of Chi recombination hot spots. We report here that both the strand that is cut and the position of the cuts relative to Chi depended on the @entity855 to @entity1025 ratio. With @entity855 in excess, Chi-dependent nicks occurred, as we have previously reported, four to six @entity2036 to the 3'-side of the Chi octamer (5'-GCTGGTGG-3') and were detected only on the strand bearing that sequence. Three differences were seen with @entity1025 in excess. 1) Chi-dependent 3'-ends were produced on the GCTGGTGG-containing strand closer to and within the Chi octamer. 2) Chi-dependent cuts occurred on the complementary DNA strand. 3) RecBCD enzyme destroyed the 3'-terminated strand of DNA from its entry point up to the vicinity of the Chi site, as others have previously reported. We show here that, with @entity1025 in excess, the enzyme continued to travel along DNA, after encountering a Chi site, releasing both strands of the DNA distal to Chi as single strands. We discuss potential biological consequences of these two modes of RecBCD enzyme-Chi interaction.
