Fast and deep phosphoproteome analysis with the Orbitrap Astral mass spectrometer.

Owing to its roles in cellular signal transduction, protein phosphorylation plays critical roles in myriad cell processes. That said, detecting and quantifying protein phosphorylation has remained a challenge. We describe the use of a novel mass spectrometer (Orbitrap Astral) coupled with data-independent acquisition (DIA) to achieve rapid and deep analysis of human and mouse phosphoproteomes.

A High Dynamic Range Ion Detector for Multireflection Time-of-Flight Analyzers.

Reflectron-based time-of-flight analyzers rely on subnanosecond detector time response to achieve acceptable resolving power for low-mid-mass, multiple-ion peaks. With the adoption of multireflection analyzers, order of magnitude longer folded ion paths relax restrictions on detector response time, allowing implementation of new technologies that greatly improve dynamic range, detector lifetime, and ion detection efficiency. A detection system is presented, integrated into a multireflection analyzer, that combines 10 keV postacceleration and focal plane correction with a unique BxE focusing, optically coupled detector, preamplification, and dual-channel digitization.

The One Hour Human Proteome.

We describe deep analysis of the human proteome in less than 1 h. We achieve this expedited proteome characterization by leveraging state-of-the-art sample preparation, chromatographic separations, and data analysis tools, and by using the new Orbitrap Astral mass spectrometer equipped with a quadrupole mass filter, a high-field Orbitrap mass analyzer, and an asymmetric track lossless (Astral) mass analyzer. The system offers high tandem mass spectrometry acquisition speed of 200 Hz and detects hundreds of peptide sequences per second within data-independent acquisition or data-dependent acquisition modes of operation.

Crowd control of ions in the Astral analyzer.

Space charge effects are the Achilles' heel of all high-resolution ion optical devices. In time-of-flight mass analyzers, these may manifest as reduction of resolving power, mass measurement shift, peak coalescence, and/or transmission losses, while highly sensitive modern ion sources and injection devices ensure that such limits are easily exceeded. Space charge effects have been investigated, by experiment and simulation study, for the astral multi-reflection analyzer, incorporating ion focusing via a pair of converging ion mirrors, and fed by a pulsed extraction ion trap.

Ultra-fast label-free quantification and comprehensive proteome coverage with narrow-window data-independent acquisition.

Mass spectrometry (MS)-based proteomics aims to characterize comprehensive proteomes in a fast and reproducible manner. Here we present the narrow-window data-independent acquisition (nDIA) strategy consisting of high-resolution MS1 scans with parallel tandem MS (MS/MS) scans of ~200 Hz using 2-Th isolation windows, dissolving the differences between data-dependent and -independent methods. This is achieved by pairing a quadrupole Orbitrap mass spectrometer with the asymmetric track lossless (Astral) analyzer which provides >200-Hz MS/MS scanning speed, high resolving power and sensitivity, and low-ppm mass accuracy.

Fast and Deep Phosphoproteome Analysis with the Orbitrap Astral Mass Spectrometer.

Owing to its roles in cellular signal transduction, protein phosphorylation plays critical roles in myriad cell processes. That said, detecting and quantifying protein phosphorylation has remained a challenge. We describe the use of a novel mass spectrometer (Orbitrap Astral) coupled with data-independent acquisition (DIA) to achieve rapid and deep analysis of human and mouse phosphoproteomes.

A Conjoined Rectilinear Collision Cell and Pulsed Extraction Ion Trap with Auxiliary DC Electrodes.

Ion traps are routinely directly coupled to mass analyzers, where they serve to suitably cool and shape an ion population prior to pulsed extraction into the analyzer proper. Such devices benefit from high duty cycle and transmission but suffer slow ion processing times caused by a compromise in the buffer gas pressure range that suitably dampens the ion kinetic energy without causing excessive scatter during extraction or within the analyzer. A rectilinear RF quadrupole ion trap has been characterized, conjoining a pressurized collision region with a pumped extraction region, and an unbroken RF interface for seamless ion transfer between them.

Parallelized Acquisition of Orbitrap and Astral Analyzers Enables High-Throughput Quantitative Analysis.

The growing trend toward high-throughput proteomics demands rapid liquid chromatography-mass spectrometry (LC-MS) cycles that limit the available time to gather the large numbers of MS/MS fragmentation spectra required for identification. Orbitrap analyzers scale performance with acquisition time and necessarily sacrifice sensitivity and resolving power to deliver higher acquisition rates. We developed a new mass spectrometer that combines a mass-resolving quadrupole, the Orbitrap, and the novel Asymmetric Track Lossless (Astral) analyzer.

Evaluating the Performance of the Astral Mass Analyzer for Quantitative Proteomics Using Data-Independent Acquisition.

We evaluate the quantitative performance of the newly released Asymmetric Track Lossless (Astral) analyzer. Using data-independent acquisition, the Thermo Scientific Orbitrap Astral mass spectrometer quantifies 5 times more peptides per unit time than state-of-the-art Thermo Scientific Orbitrap mass spectrometers, which have long been the gold standard for high-resolution quantitative proteomics. Our results demonstrate that the Orbitrap Astral mass spectrometer can produce high-quality quantitative measurements across a wide dynamic range.

Evaluating the Performance of the Astral Mass Analyzer for Quantitative Proteomics Using Data Independent Acquisition.

We evaluate the quantitative performance of the newly released Asymmetric Track Lossless (Astral) analyzer. Using data independent acquisition, the Thermo Scientific™ Orbitrap™ Astral™ mass spectrometer quantifies 5 times more peptides per unit time than state-of-the-art Thermo Scientific™ Orbitrap™ mass spectrometers, which have long been the gold standard for high resolution quantitative proteomics. Our results demonstrate that the Orbitrap Astral mass spectrometer can produce high quality quantitative measurements across a wide dynamic range.

Decoherence-Induced Universality in Simple Metal Cluster Photoelectron Angular Distributions.

Measured angular distributions of photoelectrons from size-selected copper and sodium cluster anions are demonstrated to exhibit a universal behavior independent of the initial electron state, cluster size, or material, which can be traced back to momentum conservation upon photoemission. Quantum simulations reproduce the universality under the assumption that multielectron dynamics localizes the emission on the cluster surface and renders the cluster opaque to photoelectrons, thereby quenching interference effects that would otherwise obscure this almost classical behavior.

Photoelectron imaging spectroscopy of the small sodium cluster anions Na3(-), Na5(-), and Na7(-).

We present a photoelectron imaging study of the small sodium cluster anions Na3(-), Na5(-), and Na7(-) at photon energies in the visible and near UV range (hv = 1.64-4.28 eV).

Resonances in the entrance channel of the elementary chemical reaction of fluorine and methane.

Extending the fully quantum-state-resolved description of elementary chemical reactions beyond three or four atom systems is a crucial issue in fundamental chemical research. Reactions of methane with F, Cl, H or O are key examples that have been studied prominently. In particular, reactive resonances and nonintuitive mode-selective chemistry have been reported in experimental studies for the F+CH4 →HF+CH3 reaction.

Vibrational spectroscopy of bisulfate/sulfuric acid/water clusters: structure, stability, and infrared multiple-photon dissociation intensities.

The structure and stability of mass-selected bisulfate, sulfuric acid, and water cluster anions, HSO4(-)(H2SO4)m(H2O)n, are studied by infrared photodissociation spectroscopy aided by electronic structure calculations. The triply hydrogen-bound HSO4(-)(H2SO4) configuration appears as a recurring motif in the bare clusters, while incorporation of water disrupts this stable motif for clusters with m > 1. Infrared-active vibrations predominantly involving distortions of the hydrogen-bound network are notably missing from the infrared multiple-photon dissociation (IRMPD) spectra of these ions but are fully recovered by messenger-tagging the clusters with H2.

Slow photoelectron velocity-map imaging spectroscopy of cold thiozonide (S3(-)).

We report high-resolution anion photoelectron spectra of thiozonide (S3(-)) acquired by slow electron velocity-map imaging (SEVI). The ions were cryogenically cooled within an ion trap before photodetachment. We measure an electron affinity of 2.

Slow photoelectron velocity-map imaging spectroscopy of cold negative ions.

Anion slow photoelectron velocity-map imaging (SEVI) spectroscopy is a high-resolution variant of photoelectron spectroscopy used to study the electronic and geometric structure of atoms, molecules, and clusters. To benefit from the high resolution of SEVI when it is applied to molecular species, it is essential to reduce the internal temperature of the ions as much as possible. Here, we describe an experimental setup that combines a radio-frequency ion trap to store and cool ions with the high-resolution SEVI spectrometer.

Vibrationally resolved transition state spectroscopy of the F + H2 and F + CH4 reactions.

The transition state regions of the F + para-H2, F + normal-H2, F + CH4 and F + CD4 reactions have been studied by slow electron velocity-map imaging (SEVI) spectroscopy of the anionic precursor clusters para-FH2-, normal-FH2-, FCH4 and FCD4. The F + H2 results improve on previously published photoelectron spectra, resolving a narrow peak that appears in the same position in the para-FH2 and normal-FH2- spectra, and suggesting that additional theoretical treatment is necessary to fully describe and assign the experimental results. A small peak in the para-FH2- results is also identified, matching simulations of a product resonance in the v' = 3 vibrational level.

Communication: Vibrational spectroscopy of atmospherically relevant acid cluster anions: bisulfate versus nitrate core structures.

Infrared multiple photon dissociation spectra for the smallest atmospherically relevant anions of sulfuric and nitric acid allow us to characterize structures and distinguish between clusters with a bisulfate or a nitrate core. We find that bisulfate is the main charge carrier for HSO(4)(-)·H(2)SO(4)·HNO(3) but not for NO(3)(-)·H(2)SO(4)·HNO(3). For the mixed dimer anion, we find evidence for the presence of two isomers: HSO(4)(-)·HNO(3) and NO(3)(-)·H(2)SO(4).

Ground and low-lying excited states of propadienylidene (H2C=C=C:) obtained by negative ion photoelectron spectroscopy.

A joint experimental-theoretical study has been carried out on electronic states of propadienylidene (H(2)CCC), using results from negative-ion photoelectron spectroscopy. In addition to the previously characterized X(1)A(1) electronic state, spectroscopic features are observed that belong to five additional states: the low-lying ã(3)B(1) and b(3)A(2) states, as well as two excited singlets, Ã(1)A(2) and B(1)B(1), and a higher-lying triplet, c(3)A(1). Term energies (T(0), in cm(-1)) for the excited states obtained from the data are: 10,354±11 (ã(3)B(1)); 11,950±30 (b(3)A(2)); 20,943±11 (c(3)A(1)); and 13,677±11 (Ã(1)A(2)).

Slow photoelectron velocity-map imaging spectroscopy of the phenoxide and thiophenoxide anions.

High resolution anion photodetachment spectra of the phenoxide and thiophenoxide anions were obtained with slow electron velocity-map imaging. The spectra show transitions to the X(2)B(1) neutral states of both species and to the Ã(2)B(2) state of the thiophenoxy radical. Comparison of the spectra with Franck-Condon simulations allows several gas-phase vibrations to be assigned.

Structural and electronic properties of oxidized sodium clusters: A combined photoelectron and density functional study.

Oxygen-doped sodium cluster anions Na(n)O(2) (-) with n=41-148 have been studied by low temperature photoelectron spectroscopy and density functional theory (DFT), with a particular emphasis on those sizes where a spherical electron shell closing is expected. The experimental spectra are in good agreement with the electronic density of states of the DFT lowest energy structures. The cluster structures show segregation between an ionically bonded molecular unit located at the cluster surface and a metallic part.

Premelting and postmelting in clusters.

Caloric curves for sodium clusters with N=139 and 147 atoms show a fine structure near the solid-to-liquid transition. Neither of the two sizes exhibit surface melting. For N=139, diffusion of the surface vacancies is observed, which is not possible in the closed-shell N=147 cluster.