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Identification of Multiple Kinetic Populations of DNA-Binding Proteins in Live Cells

Title data

Ho, Han N. ; Zalami, Daniel ; Köhler, Jürgen ; van Oijen, Antoine M. ; Ghodke, Harshad:
Identification of Multiple Kinetic Populations of DNA-Binding Proteins in Live Cells.
In: Biophysical Journal. Vol. 117 (2019) Issue 5 . - pp. 950-961.
ISSN 1542-0086
DOI: https://doi.org/10.1016/j.bpj.2019.07.015

Abstract in another language

Understanding how multiprotein complexes function in cells requires detailed quantitative understanding of their association and dissociation kinetics. Analysis of the heterogeneity of binding lifetimes enables the interrogation of the various intermediate states formed during the reaction. Single-molecule fluorescence imaging permits the measurement of reaction kinetics inside living organisms with minimal perturbation. However, poor photophysical properties of fluorescent probes limit the dynamic range and accuracy of measurements of off rates in live cells. Time-lapse single-molecule fluorescence imaging can partially overcome the limits of photobleaching; however, limitations of this technique remain uncharacterized. Here, we present a structured analysis of which timescales are most accessible using the time-lapse imaging approach and explore uncertainties in determining kinetic subpopulations. We demonstrate the effect of shot noise on the precision of the measurements as well as the resolution and dynamic range limits that are inherent to the method. Our work provides a convenient implementation to determine theoretical errors from measurements and to support interpretation of experimental data.

Further data

Item Type: Article in a journal
Refereed: Yes
Institutions of the University: Faculties > Faculty of Mathematics, Physics und Computer Science > Department of Physics > Chair Experimental Physics IX - Spectroscopy of Soft Matter > Chair Experimental Physics IX - Spectroscopy of Soft Matter - Univ.-Prof. Dr. Jürgen Köhler
Result of work at the UBT: Yes
DDC Subjects: 500 Science > 530 Physics
Date Deposited: 22 May 2020 06:59
Last Modified: 22 May 2020 06:59
URI: https://eref.uni-bayreuth.de/id/eprint/55197