Figure 5: The MHF complex has DNA-binding activity.
From: The structure of the FANCM–MHF complex reveals physical features for functional assembly
Calculated electrostatic on the surface of (MHF1–MHF2)2 tetramer (a) and (H3–H4)2 from nucleosome (PDB ID: 1AOI) (b). Red and blue surfaces represent negative and positive electrostatic potentials (−3.5 kBT, +3.5 kBT), respectively. The electrostatic potentials were calculated using the Adaptive Poisson-Boltzmann Solver (APBS) with PyMol APBS tools. (c) Model of (MHF1–MHF2)2 bound to DNA. Nucleosomal DNA was docked onto (MHF1–MHF2)2 tetramer through alignment of MHF1–MHF2 dimer with H2A–H2B (PDB ID: 1AOI). The left blue circle indicates the α1α1 site, the middle one for L1L2 site and the right one for MHF1 C terminus. (d) L1L2 site and αC helix would face the bound DNA modelled by alignment of MHF1 with NC2α (blue). (e) EMSA results of MHF1–MHF2 complex and the mutants. 0.6 μM DNA substrate (59 bp, Methods); proteins at lanes 1–3 (MHF1R12A/R18A/MHF2R11A/K12A): 0, 5, 10 μM, respectively; lanes 4–6 (MHF1/MHF2K27A/K29A): 5, 10, 20 μM, respectively; lanes 7–8 (MHF1K73A/R74A/MHF2): 10, 20 μM, respectively; lanes 9–10 (MHF1/MHF2): 5, 10 μM, respectively. (f) FPAs of MHF1/MHF2 and MHF1/MHF2K27A/K29A. The data of wild-type MHF was fitted according to equation (2) (Methods). (g) FPAs of mutant MHF complexes, MHF1/MHF2K27A/K29A and MHF1R110A/K111A/MHF2. (h) The DNA-binding effect of C terminus deletion on MHF1. 0.6 μM DNA substrate; proteins at lanes (MHF1/MHF2) 1–3: 0, 5, 10 μM; lanes 4–5 (MHF11−107/MHF2): 20 μM, 50 μM; lanes 6–8 (MHF11−114/MHF2): 5, 20, 50 μM, respectively.
