Development of a Platform for High-Resolution Ion Mobility Separations Coupled with Messenger Tagging Infrared Spectroscopy for High-Precision Structural Characterizations.

The ability to uniquely identify a compound requires highly precise and orthogonal measurements. Here we describe a newly developed analytical platform that integrates high resolution ion mobility and cryogenic vibrational ion spectroscopy for high-precision structural characterizations. This platform allows for the temporal separation of isomeric/isobaric ions and provides a highly sensitive description of the ion's adopted geometry in the gas phase.

Mobility Selective Ion Soft-Landing and Characterization Enabled Using Structures for Lossless Ion Manipulation.

While conventional ion-soft landing uses the mass-to-charge (/) ratio to achieve molecular selection for deposition, here we demonstrate the use of Structures for Lossless Ion Manipulation (SLIM) for mobility-based ion selection and deposition. The dynamic rerouting capabilities of SLIM were leveraged to enable the rerouting of a selected range of mobilities to a different SLIM path (rather than MS) that terminated at a deposition surface. A selected mobility range from a phosphazene ion mixture was rerouted and deposited with a current pulse (∼150 pA) resembling its mobility peak.

The Role of Ion Rotation in Ion Mobility: Ultrahigh-Precision Prediction of Ion Mobility Dependence on Ion Mass Distribution and Translational to Rotational Energy Transfer.

The role of ion rotation in determining ion mobilities is explored using the subtle gas phase ion mobility shifts based on differences in ion mass distributions between isotopomer ions that have been observed with ion mobility spectrometry (IMS) measurements. These mobility shifts become apparent for IMS resolving powers on the order of ∼1500 where relative mobilities (or alternatively momentum transfer collision cross sections; Ω) can be measured with a precision of ∼10 ppm. The isotopomer ions have identical structures and masses, differing only in their internal mass distributions, and their Ω differences cannot be predicted by widely used computational approaches, which ignore the dependence of Ω on the ion's rotational properties.

Correction: Single-conformation spectroscopy of cold, protonated PG-containing peptides: switching β-turn types and formation of a sequential type II/II' double β-turn.

Correction for 'Single-conformation spectroscopy of cold, protonated PG-containing peptides: switching β-turn types and formation of a sequential type II/II' double β-turn' by John T. Lawler , , 2022, , 2095-2109, https://doi.org/10.

Effect of Traveling Waveform Profiles on Collision Cross Section Measurements in Structures for Lossless Ion Manipulations.

We evaluated the effect of four different waveform profiles (Square, Sine, Triangle, and asymmetric Sawtooth) on the accuracy of collision cross section (CCS) measurements using traveling wave ion mobility spectrometry (TWIMS) separations in structures for lossless ion manipulations (SLIM). The effects of the waveform profiles on the accuracy of the CCS measurements were evaluated for four classes of compounds (lipids, peptides, steroids, and nucleosides) at different TW speeds (126-206 m/s) and amplitudes (15-89 V). For the lipids and peptides, the TWIMS-based CCS (CCS) deviations from the corresponding drift-tube-based CCS (CCS) measurements were significantly lower in experiments conducted using the Sawtooth waveform compared to the square waveform.

Conformer-Specific Spectroscopy and IR-Induced Isomerization of a Model γ-Peptide: Ac-γ-Phe-NHMe.

Single-conformation IR and UV spectroscopy of the prototypical capped γ-peptide Ac-γ-Phe-NHMe (γF) was carried out under jet-cooled conditions in the gas phase in order to understand its innate conformational preferences in the absence of a solvent. We obtained conformer-specific IR and UV spectra and compared the results with calculations to make assignments and explore the differences between the γ- and γ-substituted molecules. We found four conformers of γF in our experiment.

Single-conformation spectroscopy of cold, protonated PG-containing peptides: switching β-turn types and formation of a sequential type II/II' double β-turn.

D-Proline (Pro, P) is widely utilized to form β-hairpin loops in engineered peptides that would otherwise be unstructured, most often as part of a PG sub-unit that forms a β-turn. To observe whether PG facilitated this effect in short protonated peptides, conformation specific IR-UV double resonance photofragment spectra of the cold (∼10 K) protonated P and P diastereomers of the pentapeptide YAPGA was carried out in the hydride stretch (2800-3700 cm) and amide I/II (1400-1800 cm) regions. A model localized Hamiltonian was developed to better describe the 1600-1800 cm region commonly associated with the amide I vibrations.

Measurement and Theory of Gas-Phase Ion Mobility Shifts Resulting from Isotopomer Mass Distribution Changes.

The unanticipated discovery of recent ultra-high-resolution ion mobility spectrometry (IMS) measurements revealing that isotopomers─compounds that differ only in the isotopic substitution sites─can be separated has raised questions as to the physical basis for their separation. A study comparing IMS separations for two isotopomer sets in conjunction with theory and simulations accounting for ion rotational effects provides the first-ever prediction of rotation-mediated shifts. The simulations produce observable mobility shifts due to differences in gas-ion collision frequency and translational-to-rotational energy transfer.

Two-Color IRMPD Applied to Conformationally Complex Ions: Probing Cold Ion Structure and Hot Ion Unfolding.

Two-color infrared multiphoton dissociation (2C-IRMPD) spectroscopy is a technique that mitigates spectral distortions due to nonlinear absorption that is inherent to one-color IRMPD. We use a 2C-IRMPD scheme that incorporates two independently tunable IR sources, providing considerable control over the internal energy content and type of spectrum obtained by varying the trap temperature, the time delays and fluences of the two infrared lasers, and whether the first or second laser wavelength is scanned. In this work, we describe the application of this variant of 2C-IRMPD to conformationally complex peptide ions.

Digital ion trap mass analysis of high mass protein complexes using IR activation coupled with ion/ion reactions.

Native mass spectrometry (MS) focuses on measuring the masses of large biomolecular complexes and probing their structures. Large biomolecular complexes are readily introduced into mass spectrometers as gas-phase ions using electrospray ionization (ESI); however, the ions tend to be heavily adducted with solvent and salts, which leads to mass measurement errors. Various solution clean-up approaches can reduce the degree of adduction prior to introduction to the mass spectrometer.

Ultra-High-Resolution Ion Mobility Separations Over Extended Path Lengths and Mobility Ranges Achieved using a Multilevel Structures for Lossless Ion Manipulations Module.

Over the past few years, structures for lossless ion manipulations (SLIM) have used traveling waves (TWs) to move ions over long serpentine paths that can be further lengthened by routing the ions through multiple passages of the same path. Such SLIM "multipass" separations provide unprecedentedly high ion mobility resolving powers but are ultimately limited in their ion mobility range because of the range of mobilities spanned in a single pass; that is, higher mobility ions ultimately "overtake" and "lap" lower mobility ions that have experienced fewer passes, convoluting their arrival time distribution at the detector. To achieve ultrahigh resolution separations over broader mobility ranges, we have developed a new multilevel SLIM possessing multiple stacked serpentine paths.

Gas-Phase Ion/Ion Chemistry as a Probe for the Presence of Carboxylate Groups in Polypeptide Cations.

The reactivity of 1-hydroxybenzoyl triazole (HOBt) esters with the carboxylate functionality present in peptides is demonstrated in the gas phase with a doubly deprotonated dianion. The reaction forms an anhydride linkage at the carboxylate site. Upon ion trap collisional-induced dissociation (CID) of the modified peptide, the resulting spectrum shows a nominal loss of the mass of the reagent and a water molecule.

Infrared Population Transfer Spectroscopy of Cryo-Cooled Ions: Quantitative Tests of the Effects of Collisional Cooling on the Room Temperature Conformer Populations.

The single-conformation spectroscopy and infrared-induced conformational isomerization of a model protonated pentapeptide [YGPAA + H] is studied under cryo-cooled conditions in the gas phase. Building on recent results ( DeBlase , A. F.

Conformation-Specific Infrared and Ultraviolet Spectroscopy of Cold [YAPAA+H] and [YGPAA+H] Ions: A Stereochemical "Twist" on the β-Hairpin Turn.

Incorporation of the unnatural d-proline (P) stereoisomer into a polypeptide sequence is a typical strategy to encourage formation of β-hairpin loops because natural sequences are often unstructured in solution. Using conformation-specific IR and UV spectroscopy of cold (≈10 K) gas-phase ions, we probe the inherent conformational preferences of the P and P diastereomers in the protonated peptide [YAPAA+H], where only intramolecular interactions are possible. Consistent with the solution-phase studies, one of the conformers of [YAPAA+H] is folded into a charge-stabilized β-hairpin turn.

The Dimerization State of the Mammalian High Mobility Group Protein AT-Hook 2 (HMGA2).

The mammalian high mobility group protein AT-hook 2 (HMGA2) is a chromosomal architectural transcription factor involved in cell transformation and oncogenesis. It consists of three positively charged "AT-hooks" and a negatively charged C-terminus. Sequence analyses, circular dichroism experiments, and gel-filtration studies showed that HMGA2, in the native state, does not have a defined secondary or tertiary structure.