IMPROVING PEPTIDE CATABOLISM INTERPRETATION USING ION MOBILITY DATA AND SERVER-BASED DATA REVIEW WITH HELM INTEGRATION

Importance of Topic

The detailed characterization of peptide catabolism is critical for the development of peptide-based therapeutics. Understanding metabolic hot spots and degradation pathways supports design of more stable analogues, optimization of pharmacokinetics, and reduction of off-target effects. Integrating ion mobility data and standardized notation enhances the resolution and comparability of complex peptide metabolism datasets.

Objectives and Study Overview

This work presents a unified approach combining collision cross section (CCS) measurements with a server-based review platform (WebMetabase) and HELM notation to improve interpretation of peptide degradation. Eight 14-amino acid somatostatin analogues containing D-amino acids and mesityl alanine (Msa) were examined for serum stability and metabolite profiling.

Methodology and Instrumentation

• Peptide analogues were incubated in human serum at eleven time points (0 to 48 h).
• Separation and ion mobility-enabled high-resolution mass spectrometry were performed on ACQUITY UPLC coupled to Vion IMS QTof MS.
• Data processing utilized MassMeta-Site via UNIFI API, followed by upload to WebMetabase 4.0.
• HELM integration facilitated structure-level visualization of peptides and metabolites.

Main Results and Discussion

Cleavage predominantly occurred at terminal amino acids, yielding major metabolites corresponding to loss of Ala (–71 Da) and AlaGly (–128 Da). WebMetabase visualized disappearance of parent substrates and formation of key fragments. Ion mobility analysis revealed CCS shifts >3 % among –Ala metabolites, demonstrating measurable structural differences. Peptide 95 exhibited the greatest enhancement in serum stability, confirming that combined substitutions are required for substantial stabilization.

Benefits and Practical Applications

The integrated workflow enables rapid, automated processing of peptide metabolism data and remote access via web browsers. HELM notation simplifies sharing complex molecular structures. CCS tracking provides a routine metric for distinguishing isobaric analogues. This approach accelerates lead optimization and quality control in peptide drug discovery.

Future Trends and Potential Applications

• Expansion to diverse peptide scaffolds and non-standard modifications.
• Integration with AI-driven metabolite prediction tools.
• Real-time collaborative annotation and cross-site data sharing.
• Coupling with quantitative pharmacokinetic modeling for in silico design feedback loops.

Conclusion

The combination of ion mobility data, server-based review, and HELM integration offers a robust platform for detailed peptide catabolism analysis. This enables comprehensive metabolite profiling, improves structural interpretation, and supports the rational design of more stable peptide therapeutics.

Reference

1. J Kirk et al. Using MassMetaSite and WebMetabase to Process HDMSE Data Acquired on the Vion IMS QTof. Waters Technical Brief 720006362EN, 2019.
2. J Kirk et al. Characterising the Catabolism of Peptides Using Ion Mobility Enabled High Resolution MS with MassMetaSite Integration. IMSC Poster, 2018.
3. P Martín-Gago et al. Insights into Structure-Activity Relationships of Somatostatin Analogs Containing MesitylAlanine. Molecules, 2013.
4. T Radchenko et al. Software-Aided Approach to Investigate Peptide Structure and Metabolic Susceptibility. PLOS ONE, 2017.