RNA Therapeutics Institute
Genetic Phenomena | Molecular Biology
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. We benchmarked 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 used 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.
Automated Bayesian Pipeline, Single-Molecule Binding Data, TtAgo, Thermus thermophilus, DNA, Molecular Biology
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The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It is made available under a CC-BY-NC-ND 4.0 International license.
DOI of Published Version
bioRxiv 261917; doi: https://doi.org/10.1101/26191. Link to preprint on bioRxiv service.
Smith, Carlas; Jouravleva, Karina; Huisman, Maximiliaan; Jolly, Samson M.; Zamore, Phillip D.; and Grünwald, David, "An Automated Bayesian Pipeline for Rapid Analysis of Single-Molecule Binding Data" (2018). University of Massachusetts Medical School Faculty Publications. 1499.
Creative Commons License
This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 4.0 License.