Mapping dynamic protein interactions to insulin secretory granule behavior with TIRF-FRET.

ABSTRACT: Biological processes are governed by extensive networks of dynamic molecular interactions. Yet, establishing a spatial and temporal map of these interactions and their direct relationship to specific cell functions has remained a challenge. Here, we implement sensitized emission Förster resonance energy transfer (FRET) stoichiometry under total internal reflection fluorescence (TIRF) microscopy. We demonstrate through quantitative analysis and modeling that evanescent fields must be precisely matched between FRET excitation wavelengths to isolate dynamic interactions between bimolecular FRET pairs that are not entirely membrane-delimited. We then use TIRF-FRET to monitor the behavior of individual insulin-containing secretory granules at the plasma membrane of living cells, while simultane

SUBMITTER: Lam AD 

PROVIDER: S-EPMC2920663 | biostudies-literature | 2010 Aug

REPOSITORIES: biostudies-literature