Structural Basis

Only one crystal structure is available for an RdRP complex that may represent ERnNTP or *ERnNTP (Tao et al. 2002). However, the conserved nature of palm-based active sites combined with kinetic and thermodynamic analyses of site-directed mutants with nucleotide analogs has led to a structural model for nucleotide selection by 3Dpol that extrapolates well to other classes of nucleic acid polymer-ases (Gohara et al. 2004). Shown in Fig. 7 is a model for *ERnNTP (Gohara et al. 2004). The orientation of the triphosphate dictates both the stability of this complex and catalytic efficiency. The orientation of the triphosphate requires interaction with conserved structural motif A. Note that one residue of motif A, Asp-238, is located

Universal Adapted

Asp-233

Universal Adapted

Asp-233

Fig. 7 Structural basis for fidelity. The nucleotide-binding pocket of all nucleic acid polymerases with a canonical "palm"-based active site is highly conserved. The site can be divided into two parts: a region that has "universal" interactions mediated by conserved structural motif A that organize the metals and triphosphate for catalysis, and a region that has "adapted" interactions mediated by conserved structural motif B that dictate whether ribo- or 2'deoxyribonucleotides will be utilized. In the classical polymerase, there is a motif A residue located in the sugar-binding pocket capable of interacting with the motif B residue(s) involved in sugar selection. This motif A residue in other polymerases could represent the link between the nature of the bound nucleotide (correct vs incorrect) to the efficiency of nucleotidyl transfer as described herein for Asp-238 of 3Dpo1. (Gohara et al. 2004)

Fig. 7 Structural basis for fidelity. The nucleotide-binding pocket of all nucleic acid polymerases with a canonical "palm"-based active site is highly conserved. The site can be divided into two parts: a region that has "universal" interactions mediated by conserved structural motif A that organize the metals and triphosphate for catalysis, and a region that has "adapted" interactions mediated by conserved structural motif B that dictate whether ribo- or 2'deoxyribonucleotides will be utilized. In the classical polymerase, there is a motif A residue located in the sugar-binding pocket capable of interacting with the motif B residue(s) involved in sugar selection. This motif A residue in other polymerases could represent the link between the nature of the bound nucleotide (correct vs incorrect) to the efficiency of nucleotidyl transfer as described herein for Asp-238 of 3Dpo1. (Gohara et al. 2004)

in the nucleoside binding pocket. Binding of a nucleotide with an incorrect base or ribose configuration will alter the dynamics or equilibrium position of Asp-238. This perturbation will be communicated to the active site by changes in the positions of the other motif A residues, placing the triphosphate in a suboptimal orientation and leading to a destabilized *ER NTP complex with reduced catalytic efficiency.

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