Acceleration of DNA Replication of Klenow Fragment by Small Resisting Force

TL;DR

This study experimentally confirms a theoretical model of DNA replication velocity dependence on external force, showing that Klenow fragment's replication speed increases with resisting force up to a maximum, independent of assisting force.

Contribution

The paper provides the first single-molecule experimental validation of the chemomechanical coupling model for DNA polymerases, specifically for the Klenow fragment.

Findings

Replication velocity increases with resisting force up to 3.8 pN.

Velocity is independent of assisting force.

Experimental data supports the chemomechanical coupling model.

Abstract

DNA polymerases are an essential class of enzymes or molecular motors that catalyze processive DNA syntheses during DNA replications. A critical issue for DNA polymerases is their molecular mechanism of processive DNA replication. We have previously proposed a model for chemomechanical coupling of DNA polymerases, based on which the predicted results have been provided about the dependence of DNA replication velocity upon the external force on Klenow fragment of DNA polymerase I. Here, we performed single molecule measurements of the replication velocity of Klenow fragment under the external force by using magnetic tweezers. The single molecule data verified quantitatively the previous theoretical predictions, which is critical to the chemomechanical coupling mechanism of DNA polymerases. A prominent characteristic for the Klenow fragment is that the replication velocity is independent of the assisting force whereas the velocity increases largely with the increase of the resisting force, attains the maximum velocity at about 3.8 pN and then decreases with the further increase of the resisting force.