FRET-FCS detection of intralobe dynamics in calmodulin.

ABSTRACT: Fluorescence correlation spectroscopy (FCS) can be coupled with Förster resonance energy transfer (FRET) to detect intramolecular dynamics of proteins on the microsecond time scale. Here we describe application of FRET-FCS to detect fluctuations within the N-terminal and C-terminal domains of the Ca(2+)-signaling protein calmodulin. Intramolecular fluctuations were resolved by global fitting of the two fluorescence autocorrelation functions (green-green and red-red) together with the two cross-correlation functions (green-red and red-green). To match the Förster radius for FRET to the dimensions of the N-terminal and C-terminal domains, a near-infrared acceptor fluorophore (Atto 740) was coupled with a green-emitting donor (Alexa Fluor 488). Fluctuations were detected in both domains on th

SUBMITTER: Price ES 

PROVIDER: S-EPMC3151148 | biostudies-literature | 2011 Jul

REPOSITORIES: biostudies-literature