Understanding the conformational preferences of peptide ions using cold ion spectroscopy
The work presented herein focuses on using the recently constructed Instrument for Cold Ion Spectroscopy at Purdue University to investigate the conformational preferences of biologically relevant peptide ions in the gas-phase. The instrument is a custom home-built ion trap based tandem mass spectrometer that allows for the collection of UV and IR-UV double resonance spectroscopy for cryogenically cooled gas-phase ions. Recently, there has been a growing interest in ion spectroscopy as a tool for probing the intramolecular interactions and higher order structural information for biologically relevant species. The UV and IR spectra of ions can be compared to those predicated by ab initio or density function theory calculations to predict the three-dimensional structure(s) of the ions interrogated in the mass spectrometer. Specifically, the conformational preferences of peptide backbones and the resulting hydrogen bonding patterns provide critical biochemical information regarding their structure-function relationships. The primary subject of this thesis is the spectroscopic study of a prototypical protonated pentapeptide, leucine enkephalin, and its analogues. Leucine enkephalin (YGGFL) is a biologically active endogenous opioid peptide that has been extensively studied as a model peptide in mass spectrometry. Although it had been previously studied at room temperature, this work presents the first cold UV and IR-UV double resonance spectra for the ion, leading to a revised structural assignment. The assigned structure contains a single backbone conformation at vibrational temperatures of ∼10 K that is characterized by a compact hydrogen-bonding architecture in which the peptide backbone self-solvates the N-terminal ammonium group carrying the charge. In order to elucidate structural changes caused by modifying this hydrogen bonding activity, structural analogues of protonated leucine enkephalin were investigated. Because the hydrogen bonding network incorporates the charge carrier and the C-terminus, sodiated leucine enkephalin was studied to lend insight as the impact of the ammonium group and protonated C-terminally methyl esterified leucine enkephalin was studied to investigate the effects of eliminating the carboxy proton. The combined set of results provides a wealth of information regarding the inherent conformational preferences of peptide ions. Another component of this thesis is the study of the conformational effects of proline stereochemistry. Proline containing peptides are known to fragment upon collisional induced dissociation (CID) according to the “proline effect,” giving selective cleavage of the amide bond N-terminal to the Pro residue. However, it has been shown that peptides containing the non-native D-Pro enantiomer fragment through a different mechanism. Because the stereochemistry of the proline residue does not affect the gas-phase basicity of the Xxx-Pro amide bond, the differences in their fragmentation can only be attributed to differences in conformation. For this reason, two model pentatpetides, YA(L-Pro)AA and YA(D-Pro)AA, were interrogated with UV and IR-UV double resonance spectroscopy. An understanding of the conformational differences leading to the differing fragmentation pathways for these two peptides can lend insight into the origins and overall mechanism of the proline effect. Finally, the gas-phase fragmentation of a series of model synthetic foldamer based peptides was investigated using collision induced dissociation. Synthetic foldamers are oligomers that are comprised of monomer units that differ in well-defined ways from the traditional α-amino acid subunits that comprise native peptides and proteins. In particular, β-amino acids differ from their α-amino acid analogues in the addition of a single extra methylene group along the backbone. A series of three model peptides containing all β-amino acids were investigated in their protonated and alkali metal cationized forms. It was found that while protonated β-peptides fragment at the amide bond, analogous to α-peptides, metalated β-peptides fragment at the N-C β bond.
Zwier, Purdue University.
Analytical chemistry|Physical chemistry
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