IR SPECTROSCOPY ON PEPTIDES AND PROTEINS AFTER ION MOBILITY SELECTION AND IN LIQUID HELIUM DROPLETS

IR spectroscopy has become a frequently used tool to characterize gas-phase peptides and proteins. In many experiments, ions are m/z selected, irradiated by intense and tunable IR light and fragmentation is monitored as a function of IR wavelength. The presence of different conformers can, however, complicate the interpretation, as the resulting spectra represent the sum of the spectra of the individual components. We constructed a setup, in which ion mobility methods are used to obtain m/z selected ions of defined shape on which are then further investigated by IR spectroscopy. First results on peptide aggregates are presented and for some of those, the IR spectra show a transition from helical or random coil to beta sheet structures.\ In a different experiment, peptide or protein ions are captures in liquid helium droplets prior to IR spectroscopic investigation. The conditions inside a helium droplet are isothermal at 0.38 K and the interaction between the helium matrix and the molecules are weak so that only small perturbations on the molecule are expected. IR spectra for m/z selected peptides with up to 10 aminoacids and proteins containing more than 100 aminoacids have been measured. The spectra of the smaller species show resolved bands of individual oscillators, which can be used for structure assignment. For the larger species, band envelopes are obtained and for the case of highly charged proteins, a transition form helical to extended structures is observed

The role of the mobile proton in fucose migration

Fucose migration reactions represent a substantial challenge in the analysis of fucosylated glycan structures by mass spectrometry. In addition to the well-established observation of transposed fucose residues in glycan-dissociation product ions, recent experiments show that the rearrangement can also occur in intact glycan ions. These results suggest a low-energy barrier for migration of the fucose residue and broaden the relevance of fucose migration to include other types of mass spectrometry experiments, including ion mobility-mass spectrometry and ion spectroscopy. In this work, we utilize cold-ion infrared spectroscopy to provide further insight into glycan scrambling in intact glycan ions. Our results show that the mobility of the proton is a prerequisite for the migration reaction. For the prototypical fucosylated glycans Lewis x and blood group antigen H-2, the formation of adduct ions or the addition of functional groups with variable proton affinity yields significant differences in the infrared spectra. These changes correlate well with the promotion or inhibition of fucose migration through the presence or absence of a mobile proton

The structure and energetics of 3^3He and 4^4He nanodroplets doped with alkaline earth atoms

We present systematic results, based on density functional calculations, for the structure and energetics of $^3$He and $^4$He nanodroplets doped with alkaline earth atoms. We predict that alkaline earth atoms from Mg to Ba go to the center of $^3$He drops, whereas Ca, Sr, and Ba reside in a deep dimple at the surface of $^4$He drops, and Mg is at their center. For Ca and Sr, the structure of the dimples is shown to be very sensitive to the He-alkaline earth pair potentials used in the calculations. The $5s5p\leftarrow5s^2$ transition of strontium atoms attached to helium nanodroplets of either isotope has been probed in absorption experiments. The spectra show that strontium is solvated inside $^3$He nanodroplets, supporting the calculations. In the light of our findings, we emphasize the relevance of the heavier alkaline earth atoms for analyzing mixed $^3$He-$^4$He nanodroplets, and in particular, we suggest their use to experimentally probe the $^3$He-$^4$He interface.Comment: Typeset using Revtex, 20 pages and 8 Postscript file

IR action spectroscopy of glycosaminoglycan oligosaccharides

Glycosaminoglycans (GAGs) are a physio- and pharmacologically highly relevant class of complex saccharides, possessing a linear sequence and strongly acidic character. Their repetitive linear core makes them seem structurally simple at first glance, yet differences in sulfation and epimerization lead to an enormous structural diversity with only a few GAGs having been successfully characterized to date. Recent infrared action spectroscopic experiments on sulfated mono- and disaccharide ions show great promise. Here, we assess the potential of two types of gas-phase action spectroscopy approaches in the range from 1000 to 1800 cm−1 for the structural analysis of complex GAG oligosaccharides. Synthetic tetra- and pentasaccharides were chosen as model compounds for this benchmark study. Utilizing infrared multiple photon dissociation action spectroscopy at room temperature, diagnostic bands are largely unresolved. In contrast, cryogenic infrared action spectroscopy of ions trapped in helium nanodroplets yields resolved infrared spectra with diagnostic features for monosaccharide composition and sulfation pattern. The analysis of GAGs could therefore significantly benefit from expanding the conventional MS-based toolkit with gas-phase cryogenic IR spectroscopy

A selector for structural isomers of neutral molecules

We have selected and spatially separated the two conformers of 3-aminophenol (C$_6$H$_7$NO) present in a molecular beam. Analogous to the separation of ions based on their mass-to-charge ratios in a quadrupole mass filter, the neutral conformers are separated based on their different mass-to-dipole-moment ratios in an ac electric quadrupole selector. For a given ac frequency, the individual conformers experience different focusing forces, resulting in different transmissions through the selector. These experiments demonstrate that conformer-selected samples of large molecules can be prepared, offering new possibilities for the study of gas-phase biomolecules.Comment: 4 pages, 4 figures (Revtex

How Cations Change Peptide Structure

Specific interactions between cations and proteins have a strong impact on peptide and protein structure. We here shed light on the nature of the underlying interactions, especially regarding the effects on the polyamide backbone structure. To do so, we compare the conformational ensembles of model peptides in isolation and in the presence of either Li+ or Na+ cations by state-of-the-art density-functional theory (including van der Waals effects) and gas-phase infrared spectroscopy. These monovalent cations have a drastic effect on the local backbone conformation of turn-forming peptides, by disruption of the H bonding networks and the resulting severe distortion of the backbone conformations. In fact, Li+ and Na+ can even have different conformational effects on the same peptide. We also assess the predictive power of current approximate density functionals for peptide-cation systems and compare to results from established protein force fields as well as to high-level quantum chemistry (CCSD(T)).Comment: 30 pages, 7 figure

Characterization and Fate of a Septanosyl Ferrier Cation in the Gas and Solution Phases

Ferrier reactions follow a mechanistic pathway whereby Lewis acid activation of a cyclic enol ether facilitates departure of an allylic leaving group to form a glycosyl Ferrier cation. Attack on the Ferrier cation provides a new acetal linkage concurrent with the transposition of the alkene moiety. The idiosyncratic outcomes of Ferrier reactions of seven-membered ring carbohydrate-based oxepines prompted an investigation of its corresponding septanosyl Ferrier cation. Experiments that characterized the ion, including gas-phase cryogenic IR spectroscopy matched with density functional theory-calculated spectra of candidate cation structures, as well as product analysis from solution-phase Ferrier reactions, are reported here. Results from both approaches revealed an inclination of the seven-membered ring cation to contract to five-membered ring structures. Gas-phase IR spectra matched best to calculated spectra of structures in which five-membered dioxolenium formation opened the oxepine ring. In the solution phase, an attack on the ion by water led to an acyclic enal that cyclized to a C-methylene-aldehydo arabinofuranoside species. Attack by allyl trimethylsilane, on the other hand, was diastereoselective and yielded a C-allyl septanoside

Cryogenic infrared spectroscopy reveals remarkably short NH+⋯F hydrogen bonds in fluorinated phenylalanines

In past decades, hydrogen bonds involving organic fluorine have been a highly disputed topic. Obtaining clear evidence for the presence of fluorine-specific interactions is generally difficult because of their weak nature. Today, the existence of hydrogen bonds with organic fluorine is widely accepted and supported by numerous studies. However, strong bonds with short H⋯F distances remain scarce and are primarily found in designed model compounds. Using a combination of cryogenic gas-phase infrared spectroscopy and density functional theory, we here analyze a series of conformationally unrestrained fluorinated phenylalanine compounds as protonated species. The results suggest proximal NH+⋯F hydrogen bonds with an exceptionally close H⋯F distance (1.79 Å) in protonated ortho-fluorophenylalanine

Studying the Key Intermediate of RNA Autohydrolysis by Cryogenic Gas-Phase Infrared Spectroscopy

Over the course of the COVID-19 pandemic, mRNA-based vaccines have gained tremendous importance. The development and analysis of modified RNA molecules benefit from advanced mass spectrometry and require sufficient understanding of fragmentation processes. Analogous to the degradation of RNA in solution by autohydrolysis, backbone cleavage of RNA strands was equally observed in the gas phase; however, the fragmentation mechanism remained elusive. In this work, autohydrolysis-like intermediates were generated from isolated RNA dinucleotides in the gas phase and investigated using cryogenic infrared spectroscopy in helium nanodroplets. Data from both experiment and density functional theory provide evidence for the formation of a five-membered cyclic phosphate intermediate and rule out linear or six-membered structures. Furthermore, the experiments show that another prominent condensed-phase reaction of RNA nucleotides can be induced in the gas phase: the tautomerization of cytosine. Both observed reactions are therefore highly universal and intrinsic properties of the investigated molecules

Infrared Spectroscopy of Fluorenyl Cations at Cryogenic Temperatures

The notion of (anti)aromaticity is a successful concept in chemistry to explain the structure and stability of polycyclic hydrocarbons. Cyclopentadienyl and fluorenyl cations are among the most studied classical antiaromatic systems. In this work, fluorenyl cations are investigated by high-resolution gas-phase infrared spectroscopy in helium droplets. Bare fluorenyl cations are generated in the gas phase by electrospray ionization. After mass-to-charge selection, ions are captured in ultracold helium nanodroplets and probed by infrared spectroscopy using a widely tunable free-electron laser in the 600–1700 cm–1 range. The highly resolved cryogenic infrared spectra confirm, in combination with DFT computations, that all cations are present in their singlet states