, Volume 66, Issue 6, pp 1515-1545

Characterizing fluorescence recovery curves for nuclear proteins undergoing binding events

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Abstract

Fluorescence recovery after photobleaching (FRAP) is an experimental technique used to measure the mobility of proteins within the cell nucleus. After proteins of interest are fluorescently tagged for their visualization and monitoring, a small region of the nucleus is photobleached. The experimental FRAP data are obtained by recording the recovery of the fluorescence in this region over time.

In this paper, we characterize the fluorescence recovery curves for diffusing nuclear proteins undergoing binding events with an approximate spatially homogeneous structure. We analyze two mathematical models for interpreting the experimental FRAP data, namely a reaction-diffusionmodel and a compartmental model. Perturbation analysis leads to a clear explanation of two important limiting dynamical types of behavior exhibited by experimental recovery curves, namely, (1) a reduced diffusive recovery, and (2) a biphasic recovery characterized by a fast phase and a slow phase. We show how the two models, describing the same type of dynamics using different approaches, relate and share common ground. The results can be used to interpret experimental FRAP data in terms of protein dynamics and to simplify the task of parameter estimation. Application of the results is demonstrated for nuclear actin and type H1 histone.