Prerecognition Diffusion Mechanism of Human DNA Mismatch Repair Proteins along DNA: Msh2-Msh3 versus Msh2-Msh6

Abstract

DNA mismatch repair (MMR) is an important postreplication process that eliminates mispaired or unpaired nucleotides to ensure genomic replication fidelity. In humans, Msh2-Msh6 and Msh2-Msh3 are the two mismatch repair initiation factors that recognize DNA lesions. While X-ray crystal structures exist for these proteins in complex with DNA lesions, little is known about their structures during the initial search along nonspecific double-stranded DNA, because they are short-lived and difficult to determine experimentally. In this study, various computational approaches were used to sidestep these difficulties. All-atom and coarse-grained simulations based on the crystal structures of Msh2-Msh3 and Msh2-Msh6 showed no translation along the DNA, suggesting that the initial search conformation differs from the lesion-bound crystal structure. We modeled probable search-mode structures of MSH proteins and showed, using coarse-grained molecular dynamics simulations, that they can perform rotation-coupled diffusion on DNA, which is a suitable and efficient search mechanism for their function and one predicted earlier by fluorescence resonance energy transfer and fluorescence microscopy studies. This search mechanism is implemented by electrostatic interactions among the mismatch-binding domain (MBD), the clamp domains, and the DNA backbone. During simulations, their diffusion rate did not change significantly with an increasing salt concentration, which is consistent with observations from experimental studies. When the gap between their DNA-binding clamps was increased, Msh2-Msh3 diffused mostly via the clamp domains while Msh2-Msh6 still diffused using the MBD, reproducing the experimentally measured lower diffusion coefficient of Msh2-Msh6. Interestingly, Msh2-Msh3 was capable of dissociating from the DNA, whereas Msh2-Msh6 always diffused on the DNA duplex. This is consistent with the experimental observation that Msh2-Msh3, unlike Msh2-Msh6, can overcome obstacles such as nucleosomes. Our models provide a molecular picture of the different mismatch search mechanisms undertaken by Msh2-Msh6 and Msh2-Msh3, despite the similarity of their structures.

The Rad9–Rad1–Hus1 DNA Repair Clamp is Found in Microsporidia

“…MMR recognizes and corrects improperly matched DNA bases and insertions/deletions (indels) during replication, repair, and recombination processes with the help of the MutSα or MutSβ complexes, respectively ( Liu et al 2017 ). MutSα is a heterodimer composed of MSH2 and MSH6, whereas in MutSβ, MSH6 is replaced by the structural analog MSH3 ( Pal et al 2020 ). MSH2 and MSH6 were previously identified by sequence homology searches in E. cuniculi as ECU03_0540 and ECU10_0710, respectively, but no homolog of MSH3 has been identified yet.…”

Coarse-grained molecular dynamics simulations of base-pair mismatch recognition protein MutS sliding along DNA

“…Further experiments are required to prove the causal relationship. Notably, the previous CGMD simulation study of the eukaryotic MutS homolog protein, Msh2-Msh6, suggested that the sliding along DNA in the mismatch recognition conformation is slow and conformational change must be assumed to facilitate diffusion [ 21 ]. In that work, however, DNA rotation along its long axis was not modeled.…”

“…For protein/DNA interactions, we added –0.4 extra charges to them to model releases of counter ions upon protein/DNA bindings. This simple treatment of protein/DNA interactions has been successfully applied to various biological systems in our and other groups [ 21 , 32 – 37 ].…”

“…Also, ATP binding to MutS makes the polarization distribution wider, suggesting that the diffusion after the MutS conformational change is less coupled with the protein rotation [ 19 ]. Recently, the molecular dynamics (MD) simulations of human MutS homolog, Msh2-Msh6, were performed using coarse-grained (CG) models [ 21 ]. However, structural dynamics details of bacterial MutS sliding along DNA before and after ATP binding have not been addressed yet.…”

What Are the Molecular Requirements for Protein Sliding along DNA?

“…Although rotation-coupled translation was reported for many proteins, for some others it was excluded. 59 Recently, it was shown that two DNA-repair proteins with similar structures both slide at low salt concentrations, but one of them follows mostly the hopping mechanism at higher salt concentrations, 83 which might be linked to its function. The accumulated results so far thus suggest that protein structure, topology, and electrostatic potential together with the electrostatic potential of the DNA conformations 56 may modulate the balance between the usage of sliding and hopping mechanisms for the linear diffusion of proteins along DNA.…”