Universität zu Köln

Danial, John S. H., Quintana, Yuri, Ros, Uris, Shalaby, Raed, Margheritis, Eleonora G., Ramirez, Sabrina Chumpen, Ungermann, Christian, Garcia-Saez, Ana J. and Cosentino, Katia ORCID: 0000-0002-3796-3500 (2022). Systematic Assessment of the Accuracy of Subunit Counting in Biomolecular Complexes Using Automated Single-Molecule Brightness Analysis. J. Phys. Chem. Lett., 13 (3). S. 822 - 830. WASHINGTON: AMER CHEMICAL SOC. ISSN 1948-7185

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Abstract

Analysis of single-molecule brightness allows subunit counting of high-order oligomeric biomolecular complexes. Although the theory behind the method has been extensively assessed, systematic analysis of the experimental conditions required to accurately quantify the stoichiometry of biological complexes remains challenging. In this work, we develop a high-throughput, automated computational pipeline for single-molecule brightness analysis that requires minimal human input. We use this strategy to systematically quantify the accuracy of counting under a wide range of experimental conditions in simulated ground-truth data and then validate its use on experimentally obtained data. Our approach defines a set of conditions under which subunit counting by brightness analysis is designed to work optimally and helps in establishing the experimental limits in quantifying the number of subunits in a complex of interest. Finally, we combine these features into a powerful, yet simple, software that can be easily used for the analysis of the stoichiometry of such complexes.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Danial, John S. H. UNSPECIFIED UNSPECIFIED UNSPECIFIED Quintana, Yuri UNSPECIFIED UNSPECIFIED UNSPECIFIED Ros, Uris UNSPECIFIED UNSPECIFIED UNSPECIFIED Shalaby, Raed UNSPECIFIED UNSPECIFIED UNSPECIFIED Margheritis, Eleonora G. UNSPECIFIED UNSPECIFIED UNSPECIFIED Ramirez, Sabrina Chumpen UNSPECIFIED UNSPECIFIED UNSPECIFIED Ungermann, Christian UNSPECIFIED UNSPECIFIED UNSPECIFIED Garcia-Saez, Ana J. UNSPECIFIED UNSPECIFIED UNSPECIFIED Cosentino, Katia UNSPECIFIED orcid.org/0000-0002-3796-3500 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-679859
DOI:	10.1021/acs.jpclett.1c03835
Journal or Publication Title:	J. Phys. Chem. Lett.
Volume:	13
Number:	3
Page Range:	S. 822 - 830
Date:	2022
Publisher:	AMER CHEMICAL SOC
Place of Publication:	WASHINGTON
ISSN:	1948-7185
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language MICROSCOPY; PROTEINS Multiple languages Chemistry, Physical; Nanoscience & Nanotechnology; Materials Science, Multidisciplinary; Physics, Atomic, Molecular & Chemical Multiple languages
URI:	http://kups.ub.uni-koeln.de/id/eprint/67985