Cargando…
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–deoxynucleo...
Autores principales: | , , , , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Association for the Advancement of Science
2022
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8916733/ https://www.ncbi.nlm.nih.gov/pubmed/35275726 http://dx.doi.org/10.1126/sciadv.abl3522 |
_version_ | 1784668385197424640 |
---|---|
author | Turvey, Mackenzie W. Gabriel, Kristin N. Lee, Wonbae Taulbee, Jeffrey J. Kim, Joshua K. Chen, Silu Lau, Calvin J. Kattan, Rebecca E. Pham, Jenifer T. Majumdar, Sudipta Garcia, Davil Weiss, Gregory A. Collins, Philip G. |
author_facet | Turvey, Mackenzie W. Gabriel, Kristin N. Lee, Wonbae Taulbee, Jeffrey J. Kim, Joshua K. Chen, Silu Lau, Calvin J. Kattan, Rebecca E. Pham, Jenifer T. Majumdar, Sudipta Garcia, Davil Weiss, Gregory A. Collins, Philip G. |
author_sort | Turvey, Mackenzie W. |
collection | PubMed |
description | 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. |
format | Online Article Text |
id | pubmed-8916733 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | American Association for the Advancement of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-89167332022-03-21 Single-molecule Taq DNA polymerase dynamics Turvey, Mackenzie W. Gabriel, Kristin N. Lee, Wonbae Taulbee, Jeffrey J. Kim, Joshua K. Chen, Silu Lau, Calvin J. Kattan, Rebecca E. Pham, Jenifer T. Majumdar, Sudipta Garcia, Davil Weiss, Gregory A. Collins, Philip G. Sci Adv Physical and Materials Sciences 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. American Association for the Advancement of Science 2022-03-11 /pmc/articles/PMC8916733/ /pubmed/35275726 http://dx.doi.org/10.1126/sciadv.abl3522 Text en Copyright © 2022 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC). https://creativecommons.org/licenses/by-nc/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial license (https://creativecommons.org/licenses/by-nc/4.0/) , which permits use, distribution, and reproduction in any medium, so long as the resultant use is not for commercial advantage and provided the original work is properly cited. |
spellingShingle | Physical and Materials Sciences Turvey, Mackenzie W. Gabriel, Kristin N. Lee, Wonbae Taulbee, Jeffrey J. Kim, Joshua K. Chen, Silu Lau, Calvin J. Kattan, Rebecca E. Pham, Jenifer T. Majumdar, Sudipta Garcia, Davil Weiss, Gregory A. Collins, Philip G. Single-molecule Taq DNA polymerase dynamics |
title | Single-molecule Taq DNA polymerase dynamics |
title_full | Single-molecule Taq DNA polymerase dynamics |
title_fullStr | Single-molecule Taq DNA polymerase dynamics |
title_full_unstemmed | Single-molecule Taq DNA polymerase dynamics |
title_short | Single-molecule Taq DNA polymerase dynamics |
title_sort | single-molecule taq dna polymerase dynamics |
topic | Physical and Materials Sciences |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8916733/ https://www.ncbi.nlm.nih.gov/pubmed/35275726 http://dx.doi.org/10.1126/sciadv.abl3522 |
work_keys_str_mv | AT turveymackenziew singlemoleculetaqdnapolymerasedynamics AT gabrielkristinn singlemoleculetaqdnapolymerasedynamics AT leewonbae singlemoleculetaqdnapolymerasedynamics AT taulbeejeffreyj singlemoleculetaqdnapolymerasedynamics AT kimjoshuak singlemoleculetaqdnapolymerasedynamics AT chensilu singlemoleculetaqdnapolymerasedynamics AT laucalvinj singlemoleculetaqdnapolymerasedynamics AT kattanrebeccae singlemoleculetaqdnapolymerasedynamics AT phamjenifert singlemoleculetaqdnapolymerasedynamics AT majumdarsudipta singlemoleculetaqdnapolymerasedynamics AT garciadavil singlemoleculetaqdnapolymerasedynamics AT weissgregorya singlemoleculetaqdnapolymerasedynamics AT collinsphilipg singlemoleculetaqdnapolymerasedynamics |