1 research outputs found
Intrinsic Conformational Plasticity of Native EmrE Provides a Pathway for Multidrug Resistance
EmrE is a multidrug resistance efflux
pump with specificity to
a wide range of antibiotics and antiseptics. To obtain atomic-scale
insight into the attributes of the native state that encodes the broad
specificity, we used a hybrid of solution and solid-state NMR methods
in lipid bilayers and bicelles. Our results indicate that the native
EmrE dimer oscillates between inward and outward facing structural
conformations at an exchange rate (<i>k</i><sub>ex</sub>) of ∼300 s<sup>–1</sup> at 37 °C (millisecond
motions), which is ∼50-fold faster relative to the tetraÂphenylÂphosphoÂnium
(TPP<sup>+</sup>) substrate-bound form of the protein. These observables
provide quantitative evidence that the rate-limiting step in the TPP<sup>+</sup> transport cycle is not the outward–inward conformational
change in the absence of drug. In addition, using differential scanning
calorimetry, we found that the width of the gel-to-liquid crystalline
phase transition was 2 °C broader in the absence of the TPP<sup>+</sup> substrate versus its presence, which suggested that changes
in transporter dynamics can impact the phase properties of the membrane.
Interestingly, experiments with cross-linked EmrE showed that the
millisecond inward-open to outward-open dynamics was not the culprit
of the broadening. Instead, the calorimetry and NMR data supported
the conclusion that faster time scale structural dynamics (nanosecond–microsecond)
were the source and therefore impart the conformationally plastic
character of native EmrE capable of binding structurally diverse substrates.
These findings provide a clear example how differences in membrane
protein transporter structural dynamics between drug-free and bound
states can have a direct impact on the physical properties of the
lipid bilayer in an allosteric fashion