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.

中文翻译：

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单分子 Taq DNA 聚合酶动力学

Taq DNA 聚合酶在升高的温度下发挥快速构象动力学的作用——以前无法进行机械、单分子研究的机制。在这里，单壁碳纳米管晶体管记录了 Taq 分子在 22° 至 85°C 范围内处理匹配或不匹配的模板-脱氧核苷酸三磷酸对的运动。通过使用四种酶方向，核苷酸掺入的全酶闭合与 Taq 手指结构域测试互补性和方向的更快速的 20-μs 闭合区分开来。平均而言，每个核苷酸结合事件都会观察到一个短暂的关闭；即使是互补的底物对，在 72°C 下每次催化结合之间平均有 5 次瞬时关闭。瞬态关闭和催化事件的速率和持续时间几乎没有温度依赖性，