Crystal structure of Hop2-Mnd1 and mechanistic insights into its role in meiotic recombination.
Bottom Line: One end of the rod is linked to two juxtaposed winged-helix domains, and the other end is capped by extra α-helices to form a helical bundle-like structure.Deletion analysis shows that the helical bundle-like structure is sufficient for interacting with the Dmc1-ssDNA nucleofilament, and molecular modeling suggests that the curved rod could be accommodated into the helical groove of the nucleofilament.Remarkably, the winged-helix domains are juxtaposed at fixed relative orientation, and their binding to DNA is likely to perturb the base pairing according to molecular simulations.
Affiliation: Department of Biological Sciences, KAIST Institute for the Biocentury, Cancer Metastasis Control Center, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea.Show MeSH
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Mentions: In the Hop2–Mnd1 structure, the orientations of the three leucine zippers are different. Intriguingly, we found that the three leucine zippers in their respective orientations can be fitted into the helical groove in the filament of the Dmc1-ssDNA complex (Figure 6A) (44). While LZ1 and LZ2 could be snugly fitted into the groove, LZ3wCH having the capping helices appeared to cause some steric crash. In order to elaborate this observation, we produced five different deletion mutants and performed Exo I protection assays (Figure 6B). Cleavage of Dmc1-bound ssDNA by Exo I nuclease was suppressed by wild-type Hop2–Mnd1. Importantly, deletion constructs, which retain the LZ3wCH region (Figure 6B; ΔWHD, LZ2+LZ3wCH, LZ3wCH), exhibited a similar level of protection as wild type. In contrast, mutants lacking this region (Figure 6B; WHD+LZ1, ΔLZ3wCH) were far less protective. Thus, contrary to our expectation, the LZ3wCH region alone was sufficient for interacting with the Dmc1 nucleofilament. Induced-fit binding may be necessary for the interaction between LZ3wCH and the groove of the Dmc1 nucleofilament. Consistent with this notion, a proteolysis assay showed that mouse Hop2–Mnd1 complex affects the conformation of human Rad51 (45). Upon binding of LZ3wCH to the Dmc1 nucleofilament, LZ1 and LZ2 may passively position into the helical groove, and the flexibility of the LZ1–LZ2 and LZ2–LZ3 junctions may allow for their adjustments to the induced-fit conformational change of the Dmc1 nucleofilament.
Affiliation: Department of Biological Sciences, KAIST Institute for the Biocentury, Cancer Metastasis Control Center, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea.