Development of Electrostatic-to-Covalent Gas Phase Cross-linkers for Protein Structure Measurements by Mass...

Area of Science:

• Biochemistry
• Analytical Chemistry
• Structural Biology

Background:

• Native mass spectrometry (MS) enables gas-phase protein studies, but requires advanced tools for detailed structural analysis.
• Current methods may not fully exploit all potential cross-linking sites on proteins in the gas phase.

Purpose of the Study:

• To introduce and characterize a novel electrostatic-to-covalent cross-linking approach for native MS.
• To investigate the utility of this method for probing a broader range of protein residues.

Main Methods:

• Utilized gas-phase ion/ion reactions with cross-linking reagents of varying lengths.
• Cross-linked unprotonated lysine, arginine residues, and N-termini with their protonated counterparts.
• Analyzed reaction phenomenology and trends at cross-linking sites.

Main Results:

• Demonstrated that different linker lengths result in distinct cross-linking patterns.
• Enabled probing of both protonated and neutral lysine and arginine residues.
• Showcased increased accessibility to protein sites compared to native MS alone.

Conclusions:

• Electrostatic-to-covalent cross-linking is a valuable tool for native MS.
• This method provides complementary structural information to techniques like collision cross-section measurements.
• Expands the scope of residue-specific cross-linking in gas-phase protein analysis.