Erschienen:
American Association for the Advancement of Science (AAAS), 2022
Erschienen in:Science Advances
Sprache:
Englisch
DOI:
10.1126/sciadv.abl3522
ISSN:
2375-2548
Entstehung:
Anmerkungen:
Beschreibung:
<jats:p>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.</jats:p>