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Real-time dipole orientational imaging as a probe of ligand-protein interactions

journal contribution
posted on 2023-06-08, 05:53 authored by Mark Osborne
Single-molecule orientational imaging using total internal reflection fluorescence microscopy has been employed to investigate the dynamics of a protein-ligand system. Emission patterns from single tetramethylrhodamine (TMR)-biocytin molecules bound to streptavidin show that the TMR dipole adopts a limited number of favored orientations. The angular trajectories of individual dipoles exhibit remarkably similar patterns that are characteristic of single TMR molecules interacting with a relatively homogeneous population of nanoenvironments. Analysis of the polar and azimuthal angle distributions reveals a tendency for the dipole to assume three primary and two secondary orientations. Autocorrelation analysis of the dipole trajectories shows a predominantly bimodal behavior in the reorientation rates with the slow and fast components corresponding to the primary and secondary orientations, respectively. A number of mechanisms by which the observed orientations might be stabilized have been considered, in particular specific interactions between the zwitterionic TMR probe and charged residues on the streptavidin surface. Variations in the reorientation rates have been discussed in terms of local thermal fluctuations in the protein.

History

Publication status

  • Published

Journal

Journal of Physical Chemistry B

ISSN

15206106

Publisher

ACS Publications

Issue

38

Volume

109

Page range

18153-18161

Pages

9.0

Department affiliated with

  • Chemistry Publications

Notes

First demonstration of single molecule dipole imaging at video-rate with a biological application to ligand-protein interaction dynamics. Experimental design, construction, imaging, simulation and data analysis was executed by MAO who is also sole and corresponding author.

Full text available

  • No

Peer reviewed?

  • Yes

Legacy Posted Date

2012-02-06

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