Synthetic energy sensor AMPfret deciphers adenylate-dependent AMPK activation mechanism.

ABSTRACT: AMP-activated protein kinase AMPK senses and regulates cellular energy state. AMPK activation by increasing AMP and ADP concentrations involves a conformational switch within the heterotrimeric complex. This is exploited here for the construction of a synthetic sensor of cellular energetics and allosteric AMPK activation, AMPfret. Based on engineered AMPK fused to fluorescent proteins, the sensor allows direct, real-time readout of the AMPK conformational state by fluorescence resonance energy transfer (FRET). AMPfret faithfully and dynamically reports the binding of AMP and ADP to AMPK γ-CBS sites, competed by Mg²⁺-free ATP. FRET signals correlate with activation of AMPK by allosteric mechanisms and protection from dephosphorylation, attributed here to specific CBS sites, but d

SUBMITTER: Pelosse M 

PROVIDER: S-EPMC6399333 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature