Bioelectronic Measurement of Target Engagement to a Membrane-Bound Transporter

Area of Science:

• Biophysics
• Biochemistry
• Drug Discovery

Background:

• Detecting drug binding to integral membrane proteins (IMPs) is crucial for drug discovery but faces challenges with traditional methods like SPR and FRET.
• SPR's mass dependence can hinder detection of small ligands, while FRET requires potentially disruptive protein labeling.
• Integral membrane proteins (IMPs) are vital drug targets, necessitating advanced detection techniques.

Purpose of the Study:

• To introduce and validate Graphene Bio-Electronic Sensing Technology (GBEST) for label-free, mass-independent detection of ligand binding to IMPs.
• To demonstrate the capability of GBEST in a physiologically relevant, fluid lipid membrane environment.
• To overcome the limitations of existing binding assay technologies for IMP targets.

Main Methods:

• Utilizing a microfluidic chamber with a fluid lipid membrane reconstituted directly on a monolayer graphene sensor surface.
• Leveraging the electrical properties of graphene to detect minute changes in electrostatic charges caused by ligand-protein interactions.
• Employing crude membrane fractions from cells overexpressing monocarboxylate transporter 1 (MCT1) for validation.

Main Results:

• Successfully formed a fluid lipid bilayer enriched with MCT1 on the GBEST sensor.
• Obtained real-time kinetic binding data for an anti-MCT1 antibody, demonstrating mass-independent detection.
• Showcased the ability to characterize binding events in a native-like membrane environment.

Conclusions:

• GBEST provides a sensitive, label-free, and mass-independent platform for studying ligand interactions with IMPs.
• This technology enables characterization of drug target engagement in a physiologically relevant membrane context.
• GBEST holds significant potential for advancing drug discovery research targeting IMPs.