AGG JACS

Introduction

• Research Summary: Study of the structural features of AGG and its ion using IRMPD (Infrared Multiple Photon Dissociation) spectroscopy.

• Author Affiliations:

  • Sung Hwan Yoon, Julia Chamot-Rooke, Brittany R. Perkins, Amy E. Hilderbrand, John C. Poutsma, Vicki H. Wysocki from the University of Arizona, Ecole Polytechnique, and The College of William and Mary.

• Date Received: August 31, 2008.

Importance of Structural Determination

• Techniques like NMR and X-ray crystallography are benchmarks for biological structure analysis.

• Increasing use of mass spectrometry (MS) for structural investigations:

  • Soft ionization techniques (MALDI, ESI) enable the examination of biological systems from small molecules to complex proteins.

  • Mass spectrometry allows for analysis in a solvent-free environment, studying intrinsic features and interactions.

Focus on b2+ Ion Structures

• Main structures proposed for b2+ ions:

  • Diketopiperazine structure (formed by N-terminal amino group attack).

  • Oxazolone structure (formed by carbonyl oxygen attack).

• Research has investigated how ions and peptide structures can be evaluated:

  • Techniques such as MS/MS, isotope labeling, gas-phase hydrogen/deuterium exchange, ion mobility, and computational modeling.

• Past studies and indirect methods suggest that many simple peptide b2+ ions are oxazolones

• IRMPD spectroscopy: has shown potential for probing structures effectively.

Experiment Overview

• Comparison of Protonated AGG and cAG:

  • IRMPD spectra of protonated cAG (known diketopiperazine structure) revealed fragments at m/z 101 (loss of CO) and other significant ions.

  • The study used quantum chemical calculations and performed at the B3LYP/6-311++G(d,p) level to determine structures.

• Key findings in IRMPD spectrum:

  • Carbonyl stretching bands and their shifts provide structural insights.

Results from Diketopiperazine Structures

• Diketopiperazine structures protonated at amide oxygens (Gly and Ala) were found to have lower free energies than nitrogen protonated counterparts:

  • Suggests major isomers involve amide oxygen protonation.

• The protonation effect on carbonyl stretching shifts can be utilized to derive structural conclusions.

• Key Bands Observed:

  • Band at 1775 cm-1 indicates an unprotonated carbonyl amide, with shifts indicating protonation effects.

  • Presence of specific bands (1202 cm-1) indicates the stability and nature of the diketopiperazine structure.

Analysis of AGG Structure

• The b2+ ion IR spectrum revealed distinct patterns differing from cAG:

  • Blue-shifted carbonyl stretching bands at about 1970 cm-1, indicating an oxazolone structure.

  • Additional bands (1630 cm-1 and 1339 cm-1) suggest NH2 scissoring and twisting modes characteristic of oxazolones.

• As a conclusion:

  • Protonated cAG retains a diketopiperazine structure, while b2+ ion from AGG conclusively adopts an oxazolone structure.

  • This confirms previous structural hypotheses based on theoretical models and prior experiments.

Acknowledgments

• Thanks to CLIO team for assistance.

• Research supported by NIH and NSF grants, and European Commission EPITOPES project.

Supporting Information

• Availability of additional figures and spectra details to support findings.

• References provided for further exploration.