Dataset Information

An Orbitrap/Time-of-Flight Mass Spectrometer for Photofragment Ion Imaging and High-Resolution Mass Analysis of Native Macromolecular Assemblies.

ABSTRACT: We discuss the design, development, and evaluation of an Orbitrap/time-of-flight (TOF) mass spectrometry (MS)-based instrument with integrated UV photodissociation (UVPD) and time/mass-to-charge ratio (m/z)-resolved imaging for the comprehensive study of the higher-order molecular structure of macromolecular assemblies (MMAs). A bespoke TOF analyzer has been coupled to the higher-energy collisional dissociation cell of an ultrahigh mass range hybrid quadrupole-Orbitrap MS. A 193 nm excimer laser was employed to photofragment MMA ions. A combination of microchannel plates (MCPs)-Timepix (TPX) quad and MCPs-phosphor screen-TPX3CAM assemblies have been used as axial and orthogonal imaging detectors, respectively. The instrument can operate in four different modes, where the UVPD-generated fragment ions from the native MMA ions can be measured with high-mass resolution or imaged in a mass-resolved manner to reveal the relative positions of the UVPD fragments postdissociation. This information is intended to be utilized for retrieving higher-order molecular structural details that include the conformation, subunit stoichiometry, and molecular interactions as well as to understand the dissociation dynamics of the MMAs in the gas phase.

SUBMITTER: Mathew A

PROVIDER: S-EPMC10326918 | biostudies-literature | 2023 Jul

REPOSITORIES: biostudies-literature

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Publications

An Orbitrap/Time-of-Flight Mass Spectrometer for Photofragment Ion Imaging and High-Resolution Mass Analysis of Native Macromolecular Assemblies.

Mathew Anjusha A Giskes Frans F Lekkas Alexandros A Greisch Jean-François JF Eijkel Gert B GB Anthony Ian G M IGM Fort Kyle K Heck Albert J R AJR Papanastasiou Dimitris D Makarov Alexander A AA Ellis Shane R SR Heeren Ron M A RMA

Journal of the American Society for Mass Spectrometry 20230615 7

We discuss the design, development, and evaluation of an Orbitrap/time-of-flight (TOF) mass spectrometry (MS)-based instrument with integrated UV photodissociation (UVPD) and time/mass-to-charge ratio (<i>m</i>/<i>z</i>)-resolved imaging for the comprehensive study of the higher-order molecular structure of macromolecular assemblies (MMAs). A bespoke TOF analyzer has been coupled to the higher-energy collisional dissociation cell of an ultrahigh mass range hybrid quadrupole-Orbitrap MS. A 193 nm ...[more]

PMID: 37319176

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Project description:RationaleThe wide chemical diversity and complex matrices inherent to metabolomics still pose a challenge to current analytical approaches for metabolite screening. Although dedicated front-end separation techniques combined with high-resolution mass spectrometry set the benchmark from an analytical point of view, the increasing number of samples and sample complexity demand for a compromise in terms of selectivity, sensitivity and high-throughput analyses.MethodsPrior to low-field drift tube ion mobility (IM) separation and quadrupole time-of-flight mass spectrometry (QTOFMS) detection, rapid ultrahigh-performance liquid chromatography separation was used for analysis of different concentration levels of dansylated metabolites present in a yeast cell extract. For identity confirmation of metabolites at the MS2 level, an alternating frame approach was chosen and two different strategies were tested: a data-independent all-ions acquisition and a quadrupole broad band isolation (Q-BBI) directed by IM drift separation.ResultsFor Q-BBI analysis, the broad mass range isolation was successfully optimized in accordance with the distinctive drift time to m/z correlation of the dansyl derivatives. To guarantee comprehensive sampling, a broad mass isolation window of 70 Da was employed. Fragmentation was performed via collision-induced dissociation, applying a collision energy ramp optimized for the dansyl derivatives. Both approaches were studied in terms of linear dynamic range and repeatability employing ethanolic extracts of Pichia pastoris spiked with 1 μM metabolite mixture. Example data obtained for histidine and glycine showed that drift time precision (<0.01 to 0.3% RSD, n = 5) compared very well with the data reported in an earlier IM-TOFMS-based study.ConclusionsChimeric mass spectra, inherent to data-independent analysis approaches, are reduced when using a drift time directed Q-BBI approach. Additionally, an improved linear dynamic working range was observed, representing, together with a rapid front-end separation, a powerful approach for metabolite screening.

Dataset Information

An Orbitrap/Time-of-Flight Mass Spectrometer for Photofragment Ion Imaging and High-Resolution Mass Analysis of Native Macromolecular Assemblies.

Publications

An Orbitrap/Time-of-Flight Mass Spectrometer for Photofragment Ion Imaging and High-Resolution Mass Analysis of Native Macromolecular Assemblies.

Similar Datasets

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