RecJ exonuclease: substrates, products and interaction with SSB

The RecJ exonuclease from Escherichia coli degrades single-stranded DNA (ssDNA) in the 5 0 –3 0 direction and participates in homologous recombination and mismatch repair. The experiments described here address RecJ’s substrate requirements and reaction products. RecJ complexes on a variety of 5 0 single-strand tailed substrates were analyzed by electrophoretic mobility shift in the absence of Mg 2+ ion required for substrate degradation. RecJ required single-stranded tails of 7 nt or greater for robust binding; addition of Mg 2+ confirmed that substrates with 5 0 tails of 6 nt or less were poor substrates for RecJ exonuclease. RecJ is a processive exonuclease, degrading � 1000 nt after a single binding event to single-strand DNA, and releases mononucleotide products. RecJ is capable of degrading a singlestranded tail up to a double-stranded junction, although products in such reactions were heterogeneous and RecJ showed a limited ability to penetrate the duplex region. RecJ exonuclease was equally potent on 5 0 phosphorylated and unphosphorylated ends. Finally, DNA binding and nuclease activity of RecJ was specifically enhanced by the pre-addition of ssDNA-binding protein and we propose that this specific interaction may aid recruitment of RecJ.