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Cargo Loading onto Kinesin Powered Molecular Shuttles
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Linking function to global and local dynamics in an elevator-type transporter.

Didar Ciftci1,2, Chloe Martens3, Vishnu G Ghani1

  • 1Department of Physiology and Biophysics, Weill Cornell Medicine, New York, NY 10065.

Proceedings of the National Academy of Sciences of the United States of America
|December 7, 2021
PubMed
Summary
This summary is machine-generated.

Transport efficiency in membrane proteins like GltPh depends on conformational changes. We found that faster local dynamics and increased transitions boost transporter efficiency by reducing rate variations.

Keywords:
conformational dynamicsglutamate transporterhydrogen deuterium exchange mass spectrometryrate-limiting stepsingle-molecule FRET

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Area of Science:

  • Biochemistry
  • Structural Biology
  • Membrane Transport

Background:

  • Transporters facilitate molecule movement across membranes via large structural changes.
  • Understanding the kinetics of these conformational changes is crucial for transport efficiency but remains largely unknown.

Purpose of the Study:

  • To investigate the temporal relationship between structural transitions and function in the elevator-type transporter GltPh.
  • To identify molecular properties that govern transport rates and efficiency.

Main Methods:

  • Single-molecule fluorescence microscopy to track conformational changes and substrate uptake.
  • Hydrogen deuterium exchange coupled with mass spectrometry (HDX-MS) to analyze protein dynamics.

Main Results:

  • GltPh conformational changes and substrate release exhibit significant kinetic heterogeneity.
  • Mutations enhancing transition frequency and reducing substrate affinity decreased heterogeneity and improved transport efficiency.
  • HDX-MS revealed secondary structure destabilization near the substrate-binding site, correlating with increased dynamics.

Conclusions:

  • Local protein dynamics near the substrate-binding site influence global conformational changes and transport rates.
  • Increased local dynamics confer gain-of-function properties, enhancing transporter efficiency by reducing kinetic heterogeneity.