Two-Metal-Ion Catalysis: Inhibition of DNA Polymerase Activity by a Third Divalent Metal Ion.

Jimin Wang,William H Konigsberg

doi:10.3389/fmolb.2022.824794

Abstract

Almost all DNA polymerases (pols) exhibit bell-shaped activity curves as a function of both pH and Mg2+ concentration. The pol activity is reduced when the pH deviates from the optimal value. When the pH is too low the concentration of a deprotonated general base (namely, the attacking 3′-hydroxyl of the 3′ terminal residue of the primer strand) is reduced exponentially. When the pH is too high the concentration of a protonated general acid (i.e., the leaving pyrophosphate group) is reduced. Similarly, the pol activity also decreases when the concentration of the divalent metal ions deviates from its optimal value: when it is too low, the binding of the two catalytic divalent metal ions required for the full activity is incomplete, and when it is too high a third divalent metal ion binds to pyrophosphate, keeping it in the replication complex longer and serving as a substrate for pyrophosphorylysis within the complex. Currently, there is a controversy about the role of the third metal ion which we will address in this review.

Highlights

The first DNA polymerase was discovered by Arthur Kornberg and others (Bessman et al, 1958; Lehman et al, 1958), and was shown to be responsible for faithfully copying double-stranded DNA through Watson-Crick basepairing between template and primer strands and between the templating nucleotide and incoming dNTPs
ES complex for pyrophosphorylysis, its apparent inhibition with respect to the pol activity profile of the steady-state reaction is only indirect. We propose that this third Mn2+ site helps to retain the pyrophosphate product longer than usual within the ternary complex, increasing its local concentration and making pyrophosphorylysis increasingly likely
The existing biochemical and structural literature on DNA pols is fully consistent with the generalized mechanism of the two-metal-ion catalysis proposed by Thomas Steitz together with the existing transition state (TS) theory for enzymatic reactions

Summary

Introduction

The first DNA polymerase (pol) was discovered by Arthur Kornberg and others (Bessman et al, 1958; Lehman et al, 1958), and was shown to be responsible for faithfully copying double-stranded DNA through Watson-Crick basepairing between template and primer strands and between the templating nucleotide and incoming dNTPs. Third Metal Ion Inhibits Polymerases not to the enzyme itself or to the ES complex) led to the proposal of an alternate mechanism that assumes that the third divalent metal ion is directly invovled in catalysis of the chemical step of the pol reaction (Nakamura et al, 2012; Gao and Yang, 2016).

Results

Conclusion