Fluorescence anisotropy of oligomeric proteins

Article type: Research Article

Authors: Czajkowsky, Daniel M.

Affiliations: Department of Molecular Physiology and Biological Physics and Biophysics Program, University of Virginia School of Medicine, Charlottesville, VA 22908, USA Tel.: +1 434 924 1231; Fax: +1 434 982 1616; E‐mail: [email protected]

Abstract: Previous studies of protein oligomerization using time‐resolved fluorescence anisotropy assumed a single fixed probe per oligomeric complex and an identical probe orientation in complexes of different stoichiometry. However, an oligomer consisting of “n” singly labeled monomers must necessarily have “n” probes. Moreover, in the expression for the anisotropy decay, the molecular axes from which the probe orientation is defined are different for complexes that differ in stoichiometry. Here, we derive an expression for the decay of the anisotropy for molecules with any number of fixed probes, and show how an explicit understanding of the probe orientation is necessary to properly assess oligomerization.

Journal: Spectroscopy, vol. 18, no. 1, pp. 85-93, 2004