Measuring protein-ligand interactions using liquid sample desorption electrospray ionization mass spectrometry

Area of Science:

• Analytical Chemistry
• Biochemistry
• Mass Spectrometry

Background:

• Liquid sample desorption electrospray ionization-mass spectrometry (DESI-MS) previously demonstrated capability for analyzing large proteins and noncovalent complexes up to 150 kDa.
• Protein-ligand interactions are crucial in biological processes and drug development, requiring robust analytical methods for characterization.

Purpose of the Study:

• To investigate and demonstrate the utility of liquid sample DESI-MS for probing protein-ligand interactions.
• To showcase the ability of this method to rapidly determine binding stoichiometry, selectivity, and kinetics.
• To develop and validate a higher throughput method for ligand screening using liquid sample DESI-MS.

Main Methods:

• Utilized liquid sample DESI-MS to directly form intact protein-ligand complex ions by spraying ligands towards protein solutions.
• Applied a "reactive" DESI methodology for real-time analysis of binding events.
• Implemented a segmented flow approach for sequential injection of different ligands to enhance throughput for ligand screening.
• Integrated supercharging techniques to improve analyte charge without disrupting complex formation.

Main Results:

• Successfully demonstrated the formation of intact protein-ligand complex ions using liquid sample DESI-MS.
• Provided rapid insights into binding stoichiometry, selectivity, and kinetics for model systems like ribonuclease A with cytidine ligands and lysozyme with acetyl chitose ligands.
• Validated a high-throughput ligand screening method through sequential injection of segmented flows.
• Confirmed that supercharging can be effectively combined with liquid sample DESI-MS for protein-ligand interaction analysis.

Conclusions:

• Liquid sample DESI-MS is a powerful and versatile technique for the direct analysis of protein-ligand interactions.
• The "reactive" DESI approach offers rapid characterization of binding parameters and enables high-throughput screening.
• Integration with supercharging further enhances the applicability of liquid sample DESI-MS in biochemical and pharmaceutical research.