Linking function to global and local dynamics in an elevator-type transporter

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 Glt_Ph.
• 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:

• Glt_Ph 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.