Proofreading of DNA polymerase: a new kinetic model with higher-order terminal effects

Song, Yongshun; Shu, Yeqiang; Zhou, Xin; Ouyang, Zhen; Li, Ming

doi:10.1088/0953-8984/29/2/025101

Cited by 14 publications

(19 citation statements)

References 63 publications

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“…These considerations have been addressed in recent kinetic formalisms of growth velocity and fidelity during the real, non-equilibrium process, these models including nearest neighbor and higher-order effects 35–38 . In addition, within the context of information theory and thermodynamics of DNA replication, it has been recently shown that to keep the fidelity within a given tolerance, average consumed energy E must increase with the reliability of the incorporated nucleotide according the next relation 33 (see Methods):where p error is the sequence-independent probability of error per incorporated nucleotide (or error rate) in the absence of exonucleolytic proofreading and β = 1/ kT .…”

Section: Discussionmentioning

confidence: 99%

A DNA-centered explanation of the DNA polymerase translocation mechanism

Arias-González

2017

Sci Rep

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DNA polymerase couples chemical energy to translocation along a DNA template with a specific directionality while it replicates genetic information. According to single-molecule manipulation experiments, the polymerase-DNA complex can work against loads greater than 50 pN. It is not known, on the one hand, how chemical energy is transduced into mechanical motion, accounting for such large forces on sub-nanometer steps, and, on the other hand, how energy consumption in fidelity maintenance integrates in this non-equilibrium cycle. Here, we propose a translocation mechanism that points to the flexibility of the DNA, including its overstretching transition, as the principal responsible for the DNA polymerase ratcheting motion. By using thermodynamic analyses, we then find that an external load hardly affects the fidelity of the copying process and, consequently, that translocation and fidelity maintenance are loosely coupled processes. The proposed translocation mechanism is compatible with single-molecule experiments, structural data and stereochemical details of the DNA-protein complex that is formed during replication, and may be extended to RNA transcription.

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Section: Discussionmentioning

confidence: 99%

A DNA-centered explanation of the DNA polymerase translocation mechanism

Arias-González

2017

Sci Rep

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“…Its reciprocal defines the mismatch-specific fidelity which again consists of the initial discrimination and the proofreading efficiency, but the latter differs significantly from f pro defined in Sec.I, since the effective elongation rate is not but instead which includes the next-nearest neighbor effects. The same logic can be readily generalized to higher-order neighbor effects where the proof-reading efficiency will be more complicated [20, 21].…”

Section: Methodsmentioning

confidence: 99%

“…This is generally different from the F pro defined by Eq. (20). They might be approximately equal only under some conditions.…”

Section: Fig 10mentioning

confidence: 98%

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Kinetic assays of DNA polymerase fidelity: a new theoretical perspective beyond Michaelis-Menten kinetics

Shu

Ouyang

et al. 2020

Preprint

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The high fidelity of DNA polymerase is critical for the faithful replication of genomic DNA. Several approaches were proposed to quantify the fidelity of DNA polymerase. Direct measurements of the error frequency of the replication products definitely give the true fidelity but turn out very hard to implement. Two biochemical kinetic approaches, the steady-state assay and the transient-state assay, were then suggested and widely adopted. In these assays, the error frequency is indirectly estimated by using the steady-state or the transient-state kinetic theory combined with the measured kinetic rates. However, whether these indirectly estimated fidelities are equivalent to the true fidelity has never been clarified theoretically, and in particular there are different strategies to quantify the proofreading efficiency of DNAP but often lead to inconsistent results. The reason for all these confusions is that it's mathematically challenging to formulate a rigorous and general theory of the true fidelity. Recently we have succeeded to establish such a theoretical framework. In this paper, we develop this theory to make a comprehensive examination on the theoretical foundation of the kinetic assays and the relation between fidelities obtained by different methods. We conclude that while the steady-state assay and the transient-state assay can always measure the true fidelity of exonuclease-deficient DNA polymerases, they only do so for exonuclease-efficient DNA polymerases conditionally (the proper way to use these assays to quantify the proofreading efficiency is also suggested). We thus propose a new kinetic approach, the single-molecule assay, which indirectly but precisely characterizes the true fidelity of either exonuclease-deficient or exonuclease-efficient DNA polymerases.

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“…Instead, it has to be regarded as apparently reversible like Fig.1(B2). For such schemes, Michaelis-Menten kinetics is invalid, and one should use the copolymerization theory to analyze the polymerization process, such as the steady-state copolymerization theory [5,6], the first-passage (FP) method [7] developed by us and the iterated function system (IFS) developed by P.Gaspard [8] .…”

Section: Introductionmentioning

confidence: 99%

The possible fidelity-speed-proofreading cost trade-offs in DNA replication due to the exonuclease proofreading

Shu

et al. 2021

Preprint

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DNA replication is a high-fidelity information-copying processes which is realized by DNA polymerase (DNAP). The high fidelity was explained on the basis of the well-known kinetic-proofreading mechanism (KPR), under which the so-called fidelity-speed trade-off was studied theoretically. However, numerous biochemical experiments have shown that the high fidelity of DNA replication is achieved due to the initial discrimination of polymerase domain of DNAP, as well as the proofreading of the exonuclease domain of DNAP. This exonuclease-proofreading mechanism (EPR) is totally different from KPR. So the trade-off issues are worth being re-examined under EPR. In this paper, we use the first-passage method recently proposed by us to discuss the possible trade-offs in DNA replication under EPR. We show that there could be no fidelity-speed trade-off under EPR, i.e., the fidelity and the speed can be simultaneously enhanced by EPR in a large range of kinetic parameters. This provides a new perspective to understand the experimental data of the exonuclease activity of T7 DNAP and T4 DNAP. We also show that there exists the fidelity-proofreading cost trade-off, i.e., the fidelity is enhanced at the cost of increasing the futile hydrolysis of dNTP. A possible way to avoid this trade-off is to regulate the rate of DNAP translocation: slowing down the forward translocation (in the presence of the terminal mismatch) can enhance the fidelity without changing the speed and the proofreading cost. Our theoretical analysis offers deeper insights on the kinetics-function relation of DNAP.

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Proofreading of DNA polymerase: a new kinetic model with higher-order terminal effects

Cited by 14 publications

References 63 publications

A DNA-centered explanation of the DNA polymerase translocation mechanism

A DNA-centered explanation of the DNA polymerase translocation mechanism

Kinetic assays of DNA polymerase fidelity: a new theoretical perspective beyond Michaelis-Menten kinetics

The possible fidelity-speed-proofreading cost trade-offs in DNA replication due to the exonuclease proofreading

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