Cargando…

An automated Bayesian pipeline for rapid analysis of single-molecule binding data

Single-molecule binding assays enable the study of how molecular machines assemble and function. Current algorithms can identify and locate individual molecules, but require tedious manual validation of each spot. Moreover, no solution for high-throughput analysis of single-molecule binding data exi...

Descripción completa

Detalles Bibliográficos
Autores principales: Smith, Carlas S., Jouravleva, Karina, Huisman, Maximiliaan, Jolly, Samson M., Zamore, Phillip D., Grunwald, David
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6336789/
https://www.ncbi.nlm.nih.gov/pubmed/30655518
http://dx.doi.org/10.1038/s41467-018-08045-5
_version_ 1783388117691006976
author Smith, Carlas S.
Jouravleva, Karina
Huisman, Maximiliaan
Jolly, Samson M.
Zamore, Phillip D.
Grunwald, David
author_facet Smith, Carlas S.
Jouravleva, Karina
Huisman, Maximiliaan
Jolly, Samson M.
Zamore, Phillip D.
Grunwald, David
author_sort Smith, Carlas S.
collection PubMed
description Single-molecule binding assays enable the study of how molecular machines assemble and function. Current algorithms can identify and locate individual molecules, but require tedious manual validation of each spot. Moreover, no solution for high-throughput analysis of single-molecule binding data exists. Here, we describe an automated pipeline to analyze single-molecule data over a wide range of experimental conditions. In addition, our method enables state estimation on multivariate Gaussian signals. We validate our approach using simulated data, and benchmark the pipeline by measuring the binding properties of the well-studied, DNA-guided DNA endonuclease, TtAgo, an Argonaute protein from the Eubacterium Thermus thermophilus. We also use the pipeline to extend our understanding of TtAgo by measuring the protein’s binding kinetics at physiological temperatures and for target DNAs containing multiple, adjacent binding sites.
format Online
Article
Text
id pubmed-6336789
institution National Center for Biotechnology Information
language English
publishDate 2019
publisher Nature Publishing Group UK
record_format MEDLINE/PubMed
spelling pubmed-63367892019-01-22 An automated Bayesian pipeline for rapid analysis of single-molecule binding data Smith, Carlas S. Jouravleva, Karina Huisman, Maximiliaan Jolly, Samson M. Zamore, Phillip D. Grunwald, David Nat Commun Article Single-molecule binding assays enable the study of how molecular machines assemble and function. Current algorithms can identify and locate individual molecules, but require tedious manual validation of each spot. Moreover, no solution for high-throughput analysis of single-molecule binding data exists. Here, we describe an automated pipeline to analyze single-molecule data over a wide range of experimental conditions. In addition, our method enables state estimation on multivariate Gaussian signals. We validate our approach using simulated data, and benchmark the pipeline by measuring the binding properties of the well-studied, DNA-guided DNA endonuclease, TtAgo, an Argonaute protein from the Eubacterium Thermus thermophilus. We also use the pipeline to extend our understanding of TtAgo by measuring the protein’s binding kinetics at physiological temperatures and for target DNAs containing multiple, adjacent binding sites. Nature Publishing Group UK 2019-01-17 /pmc/articles/PMC6336789/ /pubmed/30655518 http://dx.doi.org/10.1038/s41467-018-08045-5 Text en © The Author(s) 2019 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
spellingShingle Article
Smith, Carlas S.
Jouravleva, Karina
Huisman, Maximiliaan
Jolly, Samson M.
Zamore, Phillip D.
Grunwald, David
An automated Bayesian pipeline for rapid analysis of single-molecule binding data
title An automated Bayesian pipeline for rapid analysis of single-molecule binding data
title_full An automated Bayesian pipeline for rapid analysis of single-molecule binding data
title_fullStr An automated Bayesian pipeline for rapid analysis of single-molecule binding data
title_full_unstemmed An automated Bayesian pipeline for rapid analysis of single-molecule binding data
title_short An automated Bayesian pipeline for rapid analysis of single-molecule binding data
title_sort automated bayesian pipeline for rapid analysis of single-molecule binding data
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6336789/
https://www.ncbi.nlm.nih.gov/pubmed/30655518
http://dx.doi.org/10.1038/s41467-018-08045-5
work_keys_str_mv AT smithcarlass anautomatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT jouravlevakarina anautomatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT huismanmaximiliaan anautomatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT jollysamsonm anautomatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT zamorephillipd anautomatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT grunwalddavid anautomatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT smithcarlass automatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT jouravlevakarina automatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT huismanmaximiliaan automatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT jollysamsonm automatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT zamorephillipd automatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata
AT grunwalddavid automatedbayesianpipelineforrapidanalysisofsinglemoleculebindingdata