Archaeal MutS5 tightly binds to Holliday junction similarly to eukaryotic MutSγ

Abstract

Archaeal DNA recombination mechanism and the related proteins are similar to those in eukaryotes. However, no functional homolog of eukaryotic MutSγ, which recognizes Holliday junction to promote homologous recombination, has been identified in archaea. Hence, the whole molecular mechanism of archaeal homologous recombination has not yet been revealed. In this study, to identify the archaeal functional homolog of MutSγ, we focused on a functionally uncharacterized MutS homolog, MutS5, from a hyperthermophilic archaeon Pyrococcus horikoshii (phMutS5). Archaeal MutS5 has a Walker ATPase motif-containing amino acid sequence that shows similarity to the ATPase domain of MutSγ. It is known that the ATPase domain of MutS homologs is also a dimerization domain. Chemical cross-linking revealed that purified phMutS5 has an ability to dimerize in solution. phMutS5 bound to Holliday junction with a higher affinity than to other branched and linear DNAs, which resembles the DNA-binding specificities of MutSγ and bacterial MutS2, a Holliday junction-resolving MutS homolog. However, phMutS5 has no nuclease activity against branched DNA unlike MutS2. The ATPase activity of phMutS5 was significantly stimulated by the presence of Holliday junction similarly to MutSγ. Furthermore, site-directed mutagenesis revealed that the ATPase activity is dependent on the Walker ATPase motif of the protein. These results suggest that archaeal MutS5 should stabilize the Holliday junction and play a role in homologous recombination, which is analogous to the function of eukaryotic MutSγ.

The GIY‐YIG endonuclease domain of Arabidopsis MutS homolog 1 specifically binds to branched DNA structures

“…One group includes bacterial MutS1 , eukaryotic MSH2, MSH3, and MSH6 ; these recognize mismatched DNA to initiate mismatch repair reaction. The other group includes bacterial MutS2 , archaeal MutS5 , eukaryotic MSH4, and MSH5 , which recognize branched DNA structures such as Holliday junction and the D‐loop structure, intermediates in homologous/homeologous recombination, to inhibit or promote the recombination.…”