Measurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces

Abstract In a high-speed single-molecule experiment with a force probe, a protein is tethered between two substrates that are manipulated to exert force on the system. To avoid nonspecific interactions between the protein and nearby substrates, the protein is usually attached to the substrates throu...

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Autores principales:	Tobias F. Bartsch, Camila M. Villasante, Felicitas E. Hengel, Ahmed Touré, Daniel M. Firester, Aaron Oswald, A. J. Hudspeth
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/3b89f7a1c30345afaf37ac9fb427dc85
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spelling	oai:doaj.org-article:3b89f7a1c30345afaf37ac9fb427dc852021-12-02T10:47:55ZMeasurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces10.1038/s41598-021-81593-x2045-2322https://doaj.org/article/3b89f7a1c30345afaf37ac9fb427dc852021-01-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-81593-xhttps://doaj.org/toc/2045-2322Abstract In a high-speed single-molecule experiment with a force probe, a protein is tethered between two substrates that are manipulated to exert force on the system. To avoid nonspecific interactions between the protein and nearby substrates, the protein is usually attached to the substrates through long, flexible linkers. This approach precludes measurements of mechanical properties with high spatial and temporal resolution, for rapidly exerted forces are dissipated into the linkers. Because mammalian hearing operates at frequencies reaching tens to hundreds of kilohertz, the mechanical processes that occur during transduction are of very short duration. Single-molecule experiments on the relevant proteins therefore cannot involve long tethers. We previously characterized the mechanical properties of protocadherin 15 (PCDH15), a protein essential for human hearing, by tethering an individual monomer through very short linkers between a probe bead held in an optical trap and a pedestal bead immobilized on a glass coverslip. Because the two confining surfaces were separated by only the length of the tethered protein, hydrodynamic coupling between those surfaces complicated the interpretation of the data. To facilitate our experiments, we characterize here the anisotropic and position-dependent diffusion coefficient of a probe in the presence of an effectively infinite wall, the coverslip, and of the immobile pedestal.Tobias F. BartschCamila M. VillasanteFelicitas E. HengelAhmed TouréDaniel M. FiresterAaron OswaldA. J. HudspethNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-7 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Tobias F. Bartsch Camila M. Villasante Felicitas E. Hengel Ahmed Touré Daniel M. Firester Aaron Oswald A. J. Hudspeth Measurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces
description	Abstract In a high-speed single-molecule experiment with a force probe, a protein is tethered between two substrates that are manipulated to exert force on the system. To avoid nonspecific interactions between the protein and nearby substrates, the protein is usually attached to the substrates through long, flexible linkers. This approach precludes measurements of mechanical properties with high spatial and temporal resolution, for rapidly exerted forces are dissipated into the linkers. Because mammalian hearing operates at frequencies reaching tens to hundreds of kilohertz, the mechanical processes that occur during transduction are of very short duration. Single-molecule experiments on the relevant proteins therefore cannot involve long tethers. We previously characterized the mechanical properties of protocadherin 15 (PCDH15), a protein essential for human hearing, by tethering an individual monomer through very short linkers between a probe bead held in an optical trap and a pedestal bead immobilized on a glass coverslip. Because the two confining surfaces were separated by only the length of the tethered protein, hydrodynamic coupling between those surfaces complicated the interpretation of the data. To facilitate our experiments, we characterize here the anisotropic and position-dependent diffusion coefficient of a probe in the presence of an effectively infinite wall, the coverslip, and of the immobile pedestal.
format	article
author	Tobias F. Bartsch Camila M. Villasante Felicitas E. Hengel Ahmed Touré Daniel M. Firester Aaron Oswald A. J. Hudspeth
author_facet	Tobias F. Bartsch Camila M. Villasante Felicitas E. Hengel Ahmed Touré Daniel M. Firester Aaron Oswald A. J. Hudspeth
author_sort	Tobias F. Bartsch
title	Measurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces
title_short	Measurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces
title_full	Measurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces
title_fullStr	Measurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces
title_full_unstemmed	Measurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces
title_sort	measurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/3b89f7a1c30345afaf37ac9fb427dc85
work_keys_str_mv	AT tobiasfbartsch measurementofhindereddiffusionincomplexgeometriesforhighspeedstudiesofsinglemoleculeforces AT camilamvillasante measurementofhindereddiffusionincomplexgeometriesforhighspeedstudiesofsinglemoleculeforces AT felicitasehengel measurementofhindereddiffusionincomplexgeometriesforhighspeedstudiesofsinglemoleculeforces AT ahmedtoure measurementofhindereddiffusionincomplexgeometriesforhighspeedstudiesofsinglemoleculeforces AT danielmfirester measurementofhindereddiffusionincomplexgeometriesforhighspeedstudiesofsinglemoleculeforces AT aaronoswald measurementofhindereddiffusionincomplexgeometriesforhighspeedstudiesofsinglemoleculeforces AT ajhudspeth measurementofhindereddiffusionincomplexgeometriesforhighspeedstudiesofsinglemoleculeforces
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Measurement of hindered diffusion in complex geometries for high-speed studies of single-molecule forces

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