ABSTRACT
The diversity of stereochemical isomers present in glycans and glycoconjugates poses a formidable challenge for comprehensive structural analysis. Typically, sophisticated mass spectrometry (MS)-based techniques are used in combination with chromatography or ion-mobility separation. However, coexisting structurally similar isomers often render an unambiguous identification impossible. Other powerful techniques such as gas-phase infrared (IR) spectroscopy have been limited to smaller glycans, since conformational flexibility and thermal activation during the measurement result in poor spectral resolution. This limitation can be overcome by using cold-ion spectroscopy. The vibrational fingerprints of cold oligosaccharide ions exhibit a wealth of well-resolved absorption features that are diagnostic for minute structural variations. The unprecedented resolution of cold-ion spectroscopy coupled with tandem MS may render this the key technology to unravel complex glycomes.
ABSTRACT
Here we present a combined experimental and theoretical study on the secondary structure of isolated proteins as a function of charge state. In infrared spectra of the proteins ubiquitin and cytochromeâ c, amideâ I (C=O stretch) and amideâ II (N-H bend) bands can be found at positions that are typical for condensed-phase proteins. For high charge states a new band appears, substantially red-shifted from the amideâ II band observed at lower charge states. The observations are interpreted in terms of Coulomb-driven transitions in secondary structures from mostly helical to extended C5 -type hydrogen-bonded structures. Support for this interpretation comes from simple energy considerations as well as from quantum chemical calculations on model peptides. This transition in secondary structure is most likely universal for isolated proteins that occur in mass spectrometric experiments.
Subject(s)
Cytochromes c/chemistry , Ubiquitin/chemistry , Hydrogen Bonding , Protein Structure, Secondary , Spectrophotometry, InfraredABSTRACT
Ultracold IR spectra of the protonated five amino acid peptide leu-enkephalin (Tyr-Gly-Gly-Phe-Leu) embedded in superfluid helium droplets have been recorded using a free-electron laser as radiation source. The results show resolved spectra, which are in good agreement with theoretical calculations, as well as with the available gas-phase data indicating that the helium environment does not induce a significant matrix-shift. In addition, the effect of the interaction between the charge and the peptide backbone has been further investigated by complexing protonated leu-enkephalin with one 18-crown-6 molecule. Good agreement between the experimental and theoretical results allow for an assignment of a preferred molecular structure.