Improved protein hydrogen/deuterium exchange mass spectrometry platform with fully automated data processing

Area of Science:

• Biochemistry
• Analytical Chemistry
• Structural Biology

Background:

• Hydrogen/deuterium exchange mass spectrometry (HDX-MS) is a standard technique for protein conformation studies.
• Existing HDX-MS methods can be prone to systematic and random errors, impacting data reliability.
• Accurate measurements are crucial for understanding protein dynamics and interactions.

Purpose of the Study:

• To describe an improved HDX-MS platform with fully automated data processing.
• To reduce systematic and random errors in HDX-MS measurements.
• To provide a robust and efficient method for analyzing protein conformational changes.

Main Methods:

• Development of an automated HDX-MS platform.
• Introduction of two types of internal standards: fast-exchanging peptides for back exchange correction and a slow-exchanging peptide (PPPI) for intrinsic rate comparison.
• Implementation of a comprehensive HDX model within MassAnalyzer software for unattended data analysis.
• Exploration of monoclonal antibody conformational transition using the platform.

Main Results:

• The platform significantly reduces errors in HDX-MS measurements.
• Automated data processing, from ion detection to HDX output, is completed within approximately 1 day.
• The system generates protection factors for backbone amide hydrogens.
• The platform successfully demonstrated the conformational transition of a monoclonal antibody.

Conclusions:

• The developed HDX-MS platform offers enhanced accuracy and automation for protein conformation studies.
• The use of internal standards effectively corrects for experimental variations.
• This automated platform provides a reliable and efficient tool for structural biology research.