Single-molecule Taq DNA polymerase dynamics.

Taq DNA polymerase functions at elevated temperatures with fast conformational dynamics-regimes previously inaccessible to mechanistic, single-molecule studies. Here, single-walled carbon nanotube transistors recorded the motions of Taq molecules processing matched or mismatched template-deoxynucleotide triphosphate pairs from 22° to 85°C. By using four enzyme orientations, the whole-enzyme closures of nucleotide incorporations were distinguished from more rapid, 20-μs closures of Taq's fingers domain testing complementarity and orientation. On average, one transient closure was observed for every nucleotide binding event; even complementary substrate pairs averaged five transient closures between each catalytic incorporation at 72°C. The rate and duration of the transient closures and the catalytic events had almost no temperature dependence, leaving all of Taq's temperature sensitivity to its rate-determining open state.

Citation

Mackenzie W Turvey, Kristin N Gabriel, Wonbae Lee, Jeffrey J Taulbee, Joshua K Kim, Silu Chen, Calvin J Lau, Rebecca E Kattan, Jenifer T Pham, Sudipta Majumdar, Davil Garcia, Gregory A Weiss, Philip G Collins. Single-molecule Taq DNA polymerase dynamics. Science advances. 2022 Mar 11;8(10):eabl3522

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PMID: 35275726

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