Atomic Resolution Insight into Sac7d Protein Binding to DNA and Associated Global Changes by Molecular Dynamics Simulations

Zacharias, Martin

doi:10.1002/ange.201900935

Cited by 2 publications

(1 citation statement)

References 30 publications

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“…Watson–Crick distance restraints were imposed on the DNA termini of the free DNA (20 kcal·mol

, allowing ±0.1 Å movement from the equilibrium bond distance) to prevent fraying of the DNA termini [ 50 ]. For the protein-DNA complexes we used slightly different distance restraints with a force constant of 2 kcal·mol

on the DNA termini, but the same as in [ 51 ], which are in accordance with B-DNA geometry. Not only were the distances between the heavy atoms of the Watson–Crick H-bonds of the DNA termini restrained but also the C1’-C1’ distance of them and the ones shifted by one base pair in the 3’ or 5’ direction (for upper and lower bounds see Supporting_Table S1 of [ 51 ]).…”

Section: Methodsmentioning

confidence: 99%

Interaction of Thymine DNA Glycosylase with Oxidised 5-Methyl-cytosines in Their Amino- and Imino-Forms

2021

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Thymine DNA Glycosylase (TDG) is an enzyme of the base excision repair mechanism and removes damaged or mispaired bases from DNA via hydrolysis of the glycosidic bond. Specificity is of high importance for such a glycosylase, so as to avoid the damage of intact DNA. Among the substrates reported for TDG are mispaired uracil and thymine but also formyl-cytosine and carboxyl-cytosine. Methyl-cytosine and hydroxylmethyl-cytosine are, in contrast, not processed by the TDG enzyme. We have in this work employed molecular dynamics simulations to explore the conformational dynamics of DNA carrying a formyl-cytosine or carboxyl-cytosine and compared those to DNA with the non-cognate bases methyl-cytosine and hydroxylmethyl-cytosine, as amino and imino tautomers. Whereas for the mispairs a wobble conformation is likely decisive for recognition, all amino tautomers of formyl-cytosine and carboxyl-cytosine exhibit the same Watson–Crick conformation, but all imino tautomers indeed form wobble pairs. The conformational dynamics of the amino tautomers in free DNA do not exhibit differences that could be exploited for recognition, and also complexation to the TDG enzyme does not induce any alteration that would indicate preferable binding to one or the other oxidised methyl-cytosine. The imino tautomers, in contrast, undergo a shift in the equilibrium between a closed and a more open, partially flipped state, towards the more open form upon complexation to the TDG enzyme. This stabilisation of the more open conformation is most pronounced for the non-cognate bases methyl-cytosine and hydroxyl-cytosine and is thus not a likely mode for recognition. Moreover, calculated binding affinities for the different forms indicate the imino forms to be less likely in the complexed DNA. These findings, together with the low probability of imino tautomers in free DNA and the indifference of the complexed amino tautomers, suggest that discrimination of the oxidised methyl-cytosines does not take place in the initial complex formation.

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“…Watson–Crick distance restraints were imposed on the DNA termini of the free DNA (20 kcal·mol

Section: Methodsmentioning

confidence: 99%

Interaction of Thymine DNA Glycosylase with Oxidised 5-Methyl-cytosines in Their Amino- and Imino-Forms

2021

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Revealing Atomic-scale Molecular Diffusion of a Plant Transcription Factor WRKY domain protein along DNA

Dai

et al. 2020

Preprint

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designed the study. Liqiang Dai conducted the MD simulations and coarse-grained simulations and analyzed the related data. Yongping Xu solved the crystal structure of WRKY-DNA complex and conducted ITC experiment. Zhenwei Du conducted the single-molecule fluorescence experiments of WRKY diffusing on DNA. AbstractTranscription factor (TF) target search on genome is highly essential for gene expression and regulation. High-resolution determination of TF diffusion along DNA remains technically challenging. Here we constructed a TF model system of the plant WRKY domain protein in complex with DNA from crystallography and demonstrated microsecond diffusion dynamics of WRKY on the DNA employing all-atom molecular dynamics (MD) simulations. Notably, we found that WRKY preferentially binds to the Crick strand of DNA with significantly stronger energetic association than to the Watson strand. The preferential binding becomes highly prominent from non-specific to specific DNA binding, but less distinct from static binding to diffusive movements of WRKY on the DNA. Remarkably, without employing acceleration forces or bias, we captured a complete one-base pair (bp) stepping cycle of WRKY tracking along major groove of DNA with homogenous (AT)n sequence, as individual protein-DNA contacts break and reform at the binding interface. Continuous tracking of WRKY forward or backward, with occasional sliding as well as strand crossing to the minor groove of DNA, have also been captured in the simulation. The processive diffusion of WRKY had been confirmed by accompanied single-molecule fluorescence assays and coarsegrained (CG) structural simulations. The study thus provides unprecedented structural dynamics details on the TF diffusion, suggests how TF possibly approaches to gene target, and supports further high-precision experimental follow-up. The stochastic movements revealed in the TF diffusion also provide general clues on how other nucleic acid walkers step and slide along DNA. Significance StatementHow transcription factors search for target genes impact on how quickly and accurately the genes are transcribed and expressed. To locate target sufficiently fast, 1D diffusion of the protein along DNA appears essential. Experimentally, it remains challenging to determine diffusional steps of protein on DNA. Here, we report all-atom equilibrium simulations of a WRKY protein binding and diffusing on DNA, revealing structural dynamics details which have not been identified previously. We unprecedently demonstrate a complete stepping cycle of the protein for one base pair on DNA within microseconds, along with stochastic stepping or sliding, directional switching, and strand crossing. Additionally, we have found preferential DNA strand association of WRKY. These suggest how protein factors approach toward target DNA sequences.

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Atomic Resolution Insight into Sac7d Protein Binding to DNA and Associated Global Changes by Molecular Dynamics Simulations

Cited by 2 publications

References 30 publications

Interaction of Thymine DNA Glycosylase with Oxidised 5-Methyl-cytosines in Their Amino- and Imino-Forms

Interaction of Thymine DNA Glycosylase with Oxidised 5-Methyl-cytosines in Their Amino- and Imino-Forms

Revealing Atomic-scale Molecular Diffusion of a Plant Transcription Factor WRKY domain protein along DNA

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