Binding of antennapedia homeodomain as studied by time-resolved fluorescence anisotropy measurements

Remo A. Hochstrasser

doi:10.1117/12.182772

17 August 1994 Binding of antennapedia homeodomain as studied by time-resolved fluorescence anisotropy measurements

Remo A. Hochstrasser

Proceedings Volume 2137, Time-Resolved Laser Spectroscopy in Biochemistry IV; (1994) https://doi.org/10.1117/12.182772
Event: OE/LASE '94, 1994, Los Angeles, CA, United States

Abstract

The accurate determination of binding constants for protein:DNA complexes is highly desirable. In the biological sciences such binding constants are often determined by gel methods that may introduce severe interactions and therefore might strongly influence the value of the physical constants. Optical methods that are also very often used are limited in protein:DNA complexes because of the overlap of the DNA and the protein absorption. To be able to use absorption or fluorescence spectroscopy it is therefore necessary to introduce labels either on the protein, the DNA, or on both. In this study an approach based on time-resolved fluorescence spectroscopy is used to determine the binding of the antennapedia homeodomain from drosophila to its operator sequence. The increase of the anisotropy decay time of fluorescently labeled DNA upon protein binding can be used to monitor binding. Oligonucleotides which include a fluorescent base analog or which are fluorescently labeled at the 5' end can be used for this purpose.

Citation Download Citation

Remo A. Hochstrasser "Binding of antennapedia homeodomain as studied by time-resolved fluorescence anisotropy measurements", Proc. SPIE 2137, Time-Resolved Laser Spectroscopy in Biochemistry IV, (17 August 1994); https://doi.org/10.1117/12.182772

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