During extension of a DNA strand using the complementary strand as a template, DNA polymerases catalyze a nucleotidyl transferase reaction in a metal dependent manner. This reaction has long been considered to require two magnesium ions, but a third active site magnesium ion was identified in recent X- and Y-family DNA polymerase product crystallographic structures. However, the function of this additional ion is not known. From our quantum mechanical/molecular mechanical calculations, we find that this additional metal ion positioned near the recently identified product metal site does not alter the reaction barrier of the chemical reaction indicating it does not participate in the catalysis of the froward reaction. Surprisingly, replacing this divalent magnesium ion by a monovalent sodium or ammonium ion significantly lowered the activation barrier. In ternary substrate complex crystallographic structures of A- and B-family DNA polymerases, a conserved lysine residue is located in the vicinity of the triphosphate of dNTP. The positively charged Nζ<sup>+</sup> of the lysine residue is found in the position occupied by the product metal identified in X- and Y-family polymerases. In fact, faster reaction rates are estimated with the replicative A- and B-family DNA polymerases. These calculations highlight the impact that an active site water network can have on the energetics of the forward reaction and how metal ions or enzyme side-chains may interact with the network to modulate the reaction barrier.

Analyzing the role of the product metal ion in DNA polymerase β catalysis

254th National Meeting (2017)

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