Dna Mismatch Repair Relies Entirely On Stochastic Transactions

DNA mismatch repair (MMR) is a highly conserved excision-resynthesis system that corrects polymerase misincorporation errors. Defects in human MMR genes cause Lynch Syndrome or hereditary non-polyposis colorectal cancer. In prototypical Escherichia coli (E. coli), cascading MutS and MutL sliding clamps initiate MMR and recruit MutH to locate and incise a hemi-methylated GATC site, which then serves as an entry point for MMR strand excision. The UvrD helicase unwinds the double-stranded DNA (dsDNA) in the direction towards the mismatch while the single-stranded DNA (ssDNA) containing the misincorporation is further degraded by an exonuclease. Single molecule imaging of the complete E. coli MMR excision process revealed that the ATP-bound MutL sliding clamp exclusively captured the UvrD helicase at the site of ssDNA-dsDNA junction, followed by tethering the UvrD to DNA junction as an unwinding processivity factor. The direction of the unwinding appeared to be random, which is determined by the side of the strand scission the MutL clamp was sliding prior to its interaction with UvrD. Moreover, the single-stranded DNA-binding protein (SSB) regulated the translocation of UvrD helicase, resulting in highly dynamic and reversible strand unwinding/rezipping tracts. The exonuclease (Exo I or Exo VII) activity was poor during strand unwinding and ultimately may be superfluous with multiple GATC incisions. These observations suggest that the traditional MMR model have missed several essential mechanical operations and may not accurately reflect MMR in vivo. In addition, our results demonstrate that MMR is governed by the stochastic nature of DNA interactions and the plasticity of protein complexes that employ thermal diffusion.