Detecting stoichiometry of macromolecular complexes in live cells using FRET.

ABSTRACT: The stoichiometry of macromolecular interactions is fundamental to cellular signalling yet challenging to detect from living cells. Fluorescence resonance energy transfer (FRET) is a powerful phenomenon for characterizing close-range interactions whereby a donor fluorophore transfers energy to a closely juxtaposed acceptor. Recognizing that FRET measured from the acceptor's perspective reports a related but distinct quantity versus the donor, we utilize the ratiometric comparison of the two to obtain the stoichiometry of a complex. Applying this principle to the long-standing controversy of calmodulin binding to ion channels, we find a surprising Ca²⁺-induced switch in calmodulin stoichiometry with Ca²⁺ channels-one calmodulin binds at basal cytosolic Ca²⁺

SUBMITTER: Ben-Johny M 

PROVIDER: S-EPMC5150656 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature