Dr Louie Slocombe

Methylation of DNA nucleobases is a naturally occurring process in living organisms. Usually, it functions as a gene regulation marker and is connected to inheritable epigenetic effects. However, the methylation of guanine in the O6 position due to external agents disrupts the hydrogen bonding between pairing bases and may have mutagenic effects. In this paper, we use density functional theory (DFT) to investigate the Double Proton Transfer (DPT) between methyl-guanine (mG) and cytosine. We compare the DPT dynamics between mG-C and unmethylated G-C using ab initio nuclear quantum dynamics as implemented in the Nuclear-Electronic Orbital (NEO-DFT) approach, where the protons involved in the transfer are described at the same quantum-mechanical level as the electrons of the system. We find that nuclear quantum effects facilitate the DPT for both systems but increase the rate of point mutations for the canonical base pair G-C more significantly. Noteworthy, when similar calculations are performed in the presence of explicit solvent and strand separation, the DPT mechanism becomes assisted by water, lowering the energy barrier of the reaction.

The adenine-thymine tautomer (A*-T*) has previously been discounted as a spontaneous mutagenesis mechanism due to the energetic instability of the tautomeric configuration. We study the stability of A*-T* while the nucleobases undergo DNA strand separation. Our calculations indicate an increase in the stability of A*-T* as the DNA strands unzip and the hydrogen bonds between the bases stretch. Molecular Dynamics simulations reveal the timescales and dynamics of DNA strand separation and the statistical ensemble of opening angles present in a biological environment. Our results demonstrate that the unwinding of DNA, an inherently out-of-equilibrium process facilitated by helicase, will change the energy landscape of the adenine-thymine tautomerisation reaction. We propose that DNA strand separation allows the stable tautomerisation of adenine-thymine, providing a feasible pathway for genetic point mutations via proton transfer between the A-T bases.

Hachimoji DNA is a synthetic nucleic acid extension of DNA, formed by an additional four bases, Z, P, S, and B, that can encode information and sustain Darwinian evolution. In this paper, we aim to look into the properties of hachimoji DNA and investigate the probability of proton transfer between the bases, resulting in base mismatch under replication. First, we present a proton transfer mechanism for hachimoji DNA, analogous to the one presented by Löwdin years prior. Then, we use density functional theory to calculate proton transfer rates, tunnelling factors and the kinetic isotope effect in hachimoji DNA. We determined that the reaction barriers are sufficiently low that proton transfer is likely to occur even at biological temperatures. Furthermore, the rates of proton transfer of hachimoji DNA are much faster than in Watson–Crick DNA due to the barrier for Z–P and S–B being 30% lower than in G–C and A–T. Suggesting that proton transfer occurs more frequently in hachimoji DNA than canonical DNA, potentially leading to a higher mutation rate. Hachimoji DNA is a synthetic nucleic acid extension of DNA formed by an additional four bases that can encode information. We examine the possibility of proton transfer between the hachimoji bases, which can result in base mismatch under replication.

Proton transfer between DNA bases can lead to mutagenic tautomers, but as their lifetimes are thought to be much shorter than DNA separation times their role during the DNA replication cycle is often overlooked. Here, the authors model the separation of the DNA base pair guanine-cytosine using density functional theory and find increased stability of the tautomer when the DNA strands unzip as they enter a helicase enzyme, effectively trapping the tautomer population. Proton transfer between the DNA bases can lead to mutagenic Guanine-Cytosine tautomers. Over the past several decades, a heated debate has emerged over the biological impact of tautomeric forms. Here, we determine that the energy required for generating tautomers radically changes during the separation of double-stranded DNA. Density Functional Theory calculations indicate that the double proton transfer in Guanine-Cytosine follows a sequential, step-like mechanism where the reaction barrier increases quasi-linearly with strand separation. These results point to increased stability of the tautomer when the DNA strands unzip as they enter the helicase, effectively trapping the tautomer population. In addition, molecular dynamics simulations indicate that the relevant strand separation time is two orders of magnitude quicker than previously thought. Our results demonstrate that the unwinding of DNA by the helicase could simultaneously slow the formation but significantly enhance the stability of tautomeric base pairs and provide a feasible pathway for spontaneous DNA mutations.

Proton transfer across hydrogen bonds in DNA can produce non-canonical nucleobase dimers and is a possible source of single-point mutations when these forms mismatch under replication. Previous computational studies have revealed this process to be energetically feasible for the guanine-cytosine (GC) base pair, but the tautomeric product (G * C *) is short-lived. In this work we reveal, for the first time, the direct effect of the replisome enzymes on proton transfer, rectifying the shortcomings of existing models. Multi-scale quantum mechanical/molecular dynamics (QM/MM) simulations reveal the effect of the bacterial PcrA Helicase on the double proton transfer in the GC base pair. It is shown that the local protein environment drastically increases the activation and reaction energies for the double proton transfer, modifying the tautomeric equilibrium. We propose a regime in which the proton transfer is dominated by tunnelling, taking place instantaneously and without atomic rearrangement of the local environment. In this paradigm, we can reconcile the metastable nature of the tautomer and show that ensemble averaging methods obscure detail in the reaction profile. Our results highlight the importance of explicit environmental models and suggest that asparagine N624 serves a secondary function of reducing spontaneous mutations in PcrA Helicase.

The misincorporation of a noncomplementary DNA base in the polymerase active site is a critical source of replication errors that can lead to genetic mutations. In this work, we model the mechanism of wobble mispairing and the subsequent rate of misincorporation errors by coupling first principles quantum chemistry calculations to an open quantum systems master equation. This methodology allows us to accurately calculate the proton transfer between bases, allowing the misincorporation and formation of mutagenic tautomeric forms of DNA bases. Our calculated rates of genetic error formation are in excellent agreement with experimental observations in DNA. Furthermore, our quantum mechanics/molecular mechanics model predicts the existence of a short-lived "tunnelling ready " configuration along the wobble reaction pathway in the polymerase active site, dramatically increasing the rate of proton transfer by a hundredfold, demonstrating that quantum tunnelling plays a critical role in determining the transcription error frequency of the polymerase.