Mass Spectrometry Experiments Research Articles

Machine learning methods for compound annotation in non-targeted mass spectrometry-A brief overview of fingerprinting, in silico fragmentation and de novo methods.

Non-targeted screenings (NTS) are essential tools in different fields, such as forensics, health and environmental sciences. NTSs often employ mass spectrometry (MS) methods due to their high throughput and sensitivity in comparison to, for example, nuclear magnetic resonance-based methods. As the identification of mass spectral signals, called annotation, is labour intensive, it has been used for developing supporting tools based on machine learning (ML). However, both the diversity of mass spectral signals and the sheer quantity of different ML tools developed for compound annotation present a challenge for researchers in maintaining a comprehensive overview of the field. In this work, we illustrate which ML-based methods are available for compound annotation in non-targeted MS experiments and provide a nuanced comparison of the ML models used in MS data analysis, unravelling their unique features and performance metrics. Through this overview we support researchers to judiciously apply these tools in their daily research. This review also offers a detailed exploration of methods and datasets to show gaps in current methods, and promising target areas, offering a starting point for developers intending to improve existing methodologies.

Hu.MAP3.0: Atlas of human protein complexes by integration of > 25,000 proteomic experiments.

Macromolecular protein complexes carry out most functions in the cell including essential functions required for cell survival. Unfortunately, we lack the subunit composition for all human protein complexes. To address this gap we integrated >25,000 mass spectrometry experiments using a machine learning approach to identify > 15,000 human protein complexes. We show our map of protein complexes is highly accurate and more comprehensive than previous maps, placing ∼75% of human proteins into their physical contexts. We globally characterize our complexes using protein co-variation data (ProteomeHD.2) and identify co-varying complexes suggesting common functional associations. Our map also generates testable functional hypotheses for 472 uncharacterized proteins which we support using AlphaFold modeling. Additionally, we use AlphaFold modeling to identify 511 mutually exclusive protein pairs in hu.MAP3.0 complexes suggesting complexes serve different functional roles depending on their subunit composition. We identify expression as the primary way cells and organisms relieve the conflict of mutually exclusive subunits. Finally, we import our complexes to EMBL-EBI's Complex Portal ( https://www.ebi.ac.uk/complexportal/home ) as well as provide complexes through our hu.MAP3.0 web interface ( https://humap3.proteincomplexes.org/ ). We expect our resource to be highly impactful to the broader research community.

Does the Formation of a Taylor Cone in a Pulsating Electrospray Directly Impact Mass Spectrometry Signals?

Electrospray ionization (ESI) remains the dominant technique in mass spectrometry (MS)-based analyses. Here, we investigated the relationship between a crucial aspect of ESI, the formation of the Taylor cone, and the MS ion current by utilizing a triple quadrupole (QqQ) mass spectrometer coupled with a streaming high-speed camera and a 3-ring electrode system. In one test, ion current over a 30-s plume gate (a ring electrode) opening was compared with the Taylor cone occurrence analyzed offline, with Spearman's correlation coefficients consistently near 0 despite parameter variations. In another test, real-time detection of Taylor cones was synchronized with QqQ-MS, selectively opening (de-energizing) the plume gate based on the Taylor cone status. This approach enabled matching the ion current with the Taylor cone occurrence. There was no apparent difference between the MS signals recorded in the presence and absence of a Taylor cone. Additionally, a Faraday plate was employed as a detector in offline experiments, revealing agreement between the frequency of liquid meniscus (Taylor cone) oscillation (∼1.92 kHz)-measured by high-speed imaging-and the frequency of spray current (∼1.93 kHz). We suggest that the lack of positive correlations in the MS experiments is due to intrinsic ion carryover during transit from the ion source to the detector and due to the insufficient data acquisition rate of the mass spectrometer, which erases short-term fluctuations of ion current.

7157 Unveiling PROP1 protein complex: NFIB as a key candidate in pituitary development

Abstract Disclosure: L.C. Iglesias García: None. M.F. Mercogliano: None. C. Schuster: None. L. Cheung: None. F. Brunello: None. V. Michan: None. M. Waldman: None. L. Defelipe: None. M. Martí: None. S.A. Camper: None. M.I. Perez-Millan: None. The transcription factor PROP1 is essential for pituitary development, and mutations in the gene are the most common cause of hypopituitarism in children. To understand PROP1 function, it is essential to identify the protein components of the PROP1 transcription complex. Rapid immunoprecipitation and mass spectrometry (RIME) experiments in GHFT1 cells, genetically engineered to express biotin-tagged PROP1, have revealed the association of PROP1 with 43 transcription factors (including β-CATENIN and the Nuclear Factor I/B). In silico analysis showed association between the armadillo domains of β-CATENIN and a predicted nuclear localization signal motif of PROP1. This interaction between PROP1 and β-CATENIN was confirmed in pituitary GHFT1 cells, where over-expression of PROP1 induces the translocation of β-CATENIN from the membrane and cytoplasm to the nucleus. Moreover, in pituitaries from Prop1df/df mutant embryos at e12.5, β-CATENIN was predominantly membrane-bound compared to wild-type embryos. These findings suggest that PROP1 is necessary for translocation of β-CATENIN to the nucleus during pituitary development.The association of PROP1 with NFIB is intriguing because PROP1 is necessary for the epithelial to mesenchymal-like transition of stem cells in pituitary development, and NFIB is associated with induction of EMT in several cancer models. We used immunohistochemistry to show that NFIB is expressed in embryonic pituitary glands from e12.5 to e16.5, and it co-localizes with the stem cell marker SOX2. To determine whether PROP1 and NFIB have common downstream targets, we performed CUT&RUN experiments for NFIB in GHFT-1 cells and compared the results with those from PROP1 ChIP-seq. Both PROP1 and NFIB bind to three sites in the NFIB gene, suggesting that NFIB is a downstream target of PROP1 and subject to autoregulation. Consistent with this idea, NFIB is reduced in pituitaries from Prop1df/df embryos at both e12.5 and e14.5 relative to normal littermates. Integration of the results from PROP1 ChIP-seq and RNA-seq (GHFT1 vs PROP1-GHFT1 cells) with NFIB CUT&RUN generated a list of 190 potential target genes. These genes are associated with cell migration, emphasizing their correlation with the EMT-like process orchestrated by PROP1 during pituitary development. In sum, we have identified an interesting set of PROP1 interacting proteins, including the transcription factor NFIB, which plays an important role in regulating stem cell differentiation in several organ systems. We have also cataloged the genome-wide set of binding sites for PROP1 and NFIB, which overlap at many genes regulating cell migration, a process that is important for pituitary stem cells to leave the niche and initiate differentiation. Presentation: 6/3/2024

Formation of Paraldehyde (C6H12O3) in Interstellar Analog Ices of Acetaldehyde Exposed to Ionizing Radiation.

Acetaldehyde (CH3CHO) plays a crucial role in the synthesis of prebiotic molecules such as amino acids, sugars, and sugar-related compounds, and in the progress of chain reaction polymerization in deep space. Here, we report the first formation of the cyclic acetaldehyde trimer - paraldehyde (C6H12O3) - in low-temperature interstellar analog ices exposed to energetic irradiation as proxies of galactic cosmic rays (GCRs). Utilizing vacuum ultraviolet photoionization reflectron time-of-flight mass spectrometry and isotopic substitution experiments, paraldehyde was identified in the gas phase during the temperature-programmed desorption of the irradiated acetaldehyde ices based on the calculated adiabatic ionization energies and isomer-specific dissociative fragmentation patterns upon photoionization. As acetaldehyde is ubiquitous throughout the interstellar medium and has been tentatively identified in interstellar ices, paraldehyde could have formed in acetaldehyde-containing ices in a cold molecular cloud and is an excellent candidate for gas-phase observation in star-forming regions via radio telescopes. The identification of paraldehyde in the gas phase from the processed acetaldehyde ices advances our understanding of how complex organic molecules can be synthesized through polymerization reactions in extraterrestrial ices exposed to GCRs.

Accounting for Digestion Enzyme Bias in Casanovo.

A key parameter of any bottom-up proteomics mass spectrometry experiment is the identity of the enzyme that is used to digest proteins in the sample into peptides. The Casanovo de novo sequencing model was trained using data that was generated with trypsin digestion; consequently, the model prefers to predict peptides that end with the amino acids "K" or "R". This bias is desirable when Casanovo is used to analyze data that was also generated using trypsin but can be problematic if the data was generated using some other digestion enzyme. In this work, we modify Casanovo to take as input the identity of the digestion enzyme alongside each observed spectrum. We then train Casanovo with data generated by using several different enzymes, and we demonstrate that the resulting model successfully learns to capture enzyme-specific behavior. However, we find, surprisingly, that this new model does not yield a significant improvement in sequencing accuracy relative to a model trained without enzyme information but using the same training set. This observation may have important implications for future attempts to make use of experimental metadata in de novo sequencing models.

Complexation of Diserinol Isophthalamide with Phosphorylated Biomolecules in Electrospray Ionization Mass Spectrometry

Electrospray ionization (ESI) enables gentle transfer of biomolecules from solution to vacuum, facilitating the study of biomolecular structure under highly controlled conditions. However, biomolecules are desolvated during the ESI process, and the loss of ionic hydrogen bonds to solvent molecules can drive structural rearrangement, most prominently at solvent-exposed charge sites. Microsolvation reagents can bind to these bare charge sites in ESI mass spectrometry (ESI–MS) experiments, providing alternative intermolecular interaction partners. Previously, 18-crown-6 was shown to be an effective reagent for binding to cationic monoalkylammonium residues. More recently, diserinol isophthalamide (DIP) was reported as an analogous anionic microsolvation reagent, primarily for carboxylate residues of small model peptides. Herein, we expand upon this work to examine the complexation of DIP, 1,1’-(1,2-phenylene)bis(3-phenylurea) (PBP), and triclocarban (TCC) with molecules featuring a terminal or linking phosphate moiety. Specifically, using ESI–MS, we assess the binding of these reagents with dimethyl phosphate (DMP), cyclic adenosine monophosphate (cAMP), dibutyryl cAMP, RNA dinucleotides ApU and CpG, and angiotensin II phosphate (DRVpYIHPF). For DMP, the smallest target molecule, reagents TCC, PBP and DIP showed favorable adduction. However, for larger systems, PBP and TCC showed reduced complexation, which was attributed to steric hindrance from the terminal aromatic moieties of PBP and the limited hydrogen bonding network of TCC. Overall, of the three reagents, DIP showed the most consistent performance for anionic microsolvation of phosphate groups, facilitating future studies of gas-phase biomolecular structure and the effects of microsolvation.

Meta-substituted thienoviologen with enhanced radical stability via π-π interaction modulation for neutral aqueous organic flow batteries

The aqueous organic redox flow batteries (AORFBs) are recognized as the most promising large-scale storage technology for long-duration energy storage (LDES). Although viologen derivatives are widely used as anolyte materials for AORFBs, their practical application is strongly limited by weak conjugation and unstable radicals. Here, we present a novel thienoviologen derivative, [(NPr)2SV]Cl4, achieved utilizing a meta-substitution approach based on the pristine viologen derivatives ( [(NPr)2V]Cl4). Compared to other ortho- and para-substitution methods, this strategy features a locked plane configuration (0.14°), effectively regulating π-π interactions and suppressing reactivity between radical and oxygen, which is confirmed via X-ray single-crystal structural analyses, spectroscopy techniques, molecular dynamics simulation accompanied by linear ion trap mass spectrometry experiments. Additionally, the meta-substituted [(NPr)2SV]Cl4 significantly improves water solubility (2.39 M), enhances aromaticity, and extends radical lifetime compared with para-substituted [(NPr)2TV]Cl4. Consequently, the cycling stability of the [(NPr)2SV]Cl4-based AORFB over 2500 cycles is 4.1 times higher than that of [(NPr)2TV]Cl4. Furthermore, the 0.5 M battery delivers an impressive 99.83% capacity retention during 200 cycles and a power density of 147 mW cm-2.

LncOCMRL1 promotes oral squamous cell carcinoma growth and metastasis via the RRM2/EMT pathway

BackgroundLong noncoding RNAs (lncRNAs) are widely involved in cancer development and progression, but the functions of most lncRNAs have not yet been elucidated. Metastasis is the main factor restricting the therapeutic outcomes of various cancer types, including oral squamous cell carcinoma (OSCC). Therefore, exploring the key lncRNAs that regulate OSCC metastasis and elucidating their molecular mechanisms will facilitate the development of new strategies for effective OSCC therapy.MethodsWe analyzed the lncRNA expression profiles of tumor tissues from OSCC patients with and without cervical lymph node metastasis, and OSCC cell lines. We revealed high expression of oral squamous cell carcinoma metastasis-related lncRNA 1 (lncOCMRL1) in OSCC patient tumor tissues with lymph node metastasis and highly metastatic OSCC cell lines. The effects of lncOCMRL1 knockdown on the invasion, migration and proliferation abilities of OSCC cells were explored through qRT-PCR, Transwell, colony formation, and cell proliferation experiments. The mechanism by which lncOCMRL1 promotes OSCC metastasis and proliferation was explored through RNA pull-down, silver staining, mass spectrometry, RIP, and WB experiments. To increase its translational potential, we developed a reduction-responsive nanodelivery system to deliver siRNA for antitumor therapy.ResultsWe determined that lncOCMRL1 is highly expressed in OSCC metastatic tumor tissues and cells. Functional studies have shown that high lncOCMRL1 expression can promote the growth and metastasis of OSCC cells both in vivo and in vitro. Mechanistically, lncOCMRL1 could induce epithelial-mesenchymal transition (EMT) via the suppression of RRM2 ubiquitination and thereby promote the proliferation, invasion, and migration of OSCC cells. We further constructed reduction-responsive nanoparticles (NPs) for the systemic delivery of siRNAs targeting lncOCMRL1 and demonstrated their high efficacy in silencing lncOCMRL1 expression in vivo and significantly inhibited OSCC tumor growth and metastasis.ConclusionsOur results suggest that lncOCMRL1 is a reliable target for blocking lymph node metastasis in OSCC.

Mycobacteria that cause tuberculosis have retained ancestrally acquired genes for the biosynthesis of chemically diverse terpene nucleosides.

Mycobacterium tuberculosis (Mtb) releases the unusual terpene nucleoside 1-tuberculosinyladenosine (1-TbAd) to block lysosomal function and promote survival in human macrophages. Using conventional approaches, we found that genes Rv3377c and Rv3378c, but not Rv3376, were necessary for 1-TbAd biosynthesis. Here, we introduce linear models for mass spectrometry (limms) software as a next-generation lipidomics tool to study the essential functions of lipid biosynthetic enzymes on a whole-cell basis. Using limms, whole-cell lipid profiles deepened the phenotypic landscape of comparative mass spectrometry experiments and identified a large family of approximately 100 terpene nucleoside metabolites downstream of Rv3378c. We validated the identity of previously unknown adenine-, adenosine-, and lipid-modified tuberculosinol-containing molecules using synthetic chemistry and collisional mass spectrometry, including comprehensive profiling of bacterial lipids that fragment to adenine. We tracked terpene nucleoside genotypes and lipid phenotypes among Mycobacterium tuberculosis complex (MTC) species that did or did not evolve to productively infect either human or nonhuman mammals. Although 1-TbAd biosynthesis genes were thought to be restricted to the MTC, we identified the locus in unexpected species outside the MTC. Sequence analysis of the locus showed nucleotide usage characteristic of plasmids from plant-associated bacteria, clarifying the origin and timing of horizontal gene transfer to a pre-MTC progenitor. The data demonstrated correlation between high level terpene nucleoside biosynthesis and mycobacterial competence for human infection, and 2 mechanisms of 1-TbAd biosynthesis loss. Overall, the selective gain and evolutionary retention of tuberculosinyl metabolites in modern species that cause human TB suggest a role in human TB disease, and the newly discovered molecules represent candidate disease-specific biomarkers.

A ratiometric fluorescent probe based on phenothiazine and quinazolinone for rapid detection of ClO− and its application

A novel fluorescent probe (BPQ-1) with large conjugated system (phenothiazine was conjugated with quinazolinone) was synthesized. BPQ-1 showed a remarkable ratiometric fluorescence response to ClO− within 10 s, and achieved large stokes shift (168 nm), high sensitivity (LOD = 16.7 nM) and excellent selectivity. Among the analytes to be detected, there is a significant change from bright yellow fluorescence to cyan fluorescence only when ClO− is added to BPQ-1. Through density functional theory calculation and mass spectrometry experiments, the fluorescence response mechanisms of the BPQ-1 are given. In addition, BPQ-1 has been successfully applied to the detection of ClO− in real water samples and test strips for detecting ClO− with different concentrations have been prepared. In particular, the probe has been successfully applied to the fluorescence imaging of exogenous ClO− in living cells.

Sensing signal augmentation by flow rate modulation of carrier gas for accurate differentiation of complex odours

ABSTRACT For olfactory sensors, clear differentiation of complex odour samples requires diverse information. To obtain such information, hardware modifications, such as introducing additional channels with different physical/chemical properties, are usually needed. In this study, we present a new approach to augmenting the sensing signals of an olfactory sensor by modulating the flow rate of the carrier gas. The headspace vapour of complex odours is measured using a sensing system of nanomechanical sensor (Membrane-type Surface stress Sensor, MSS). The resulting data set is quantitatively evaluated using the Davies-Bouldin index (DBI) of principal component analysis (PCA). The increasing number of sensing signals obtained at different gas flow rates leads to a decrease in the DBI, achieving better cluster separation between different odours. Such gas flow effects can be attributed to several factors, including the sample evaporation and the equilibrium of the gas-liquid and gas-solid interfaces. Proton-transfer-reaction time-of-flight mass spectrometry (PTR-TOF-MS) experiments reveal that the compositions of odour samples vary with the different gas flow rates. It is demonstrated that a simple technique for modulating gas flow rates can significantly improve the differentiation performance of complex odours, providing an additional degree of freedom in olfactory sensing.

Hydrogen Evolution Reactions of Hydrocarbons and Hydroborons Promoted by Superatomic Nbn- Clusters.

Hydrogen evolution reactions (HER) are crucial for producing renewable and ecological hydrogen energy. Here we report the finding that ethyl acetylene (C2H2), borane (B2H6), and benzene (C6H6) undergo drastic dehydrogenation reactions upon interaction with niobium cluster anions (Nbn-). This finding was enabled by our very sensitive and specialized mass spectrometer, which monitored the cluster ions and the resulting metal carbide and boride products in real time. Through mass spectrometry experiments and density functional theory computations, we delved into the varying reactivities of these common gas molecules with small Nbn- clusters and elucidated the underlying reaction mechanisms. These findings shed light on the HER mechanisms of hydrocarbon and hydroboron molecules on superatomic Nbn- clusters, furnishing valuable insights pertinent to the advancement of hydrogen energy resources.

Activation of parkin by a molecular glue

Mutations in parkin and PINK1 cause early-onset Parkinson’s disease (EOPD). The ubiquitin ligase parkin is recruited to damaged mitochondria and activated by PINK1, a kinase that phosphorylates ubiquitin and the ubiquitin-like domain of parkin. Activated phospho-parkin then ubiquitinates mitochondrial proteins to target the damaged organelle for degradation. Here, we present the mechanism of activation of a new class of small molecule allosteric modulators that enhance parkin activity. The compounds act as molecular glues to enhance the ability of phospho-ubiquitin (pUb) to activate parkin. Ubiquitination assays and isothermal titration calorimetry with the most active compound (BIO-2007817) identify the mechanism of action. We present the crystal structure of a closely related compound (BIO-1975900) bound to a complex of parkin and two pUb molecules. The compound binds next to pUb on RING0 and contacts both proteins. Hydrogen-deuterium exchange mass spectrometry (HDX-MS) experiments confirm that activation occurs through release of the catalytic Rcat domain. In organello and mitophagy assays demonstrate that BIO-2007817 partially rescues the activity of parkin EOPD mutants, R42P and V56E, offering a basis for the design of activators as therapeutics for Parkinson’s disease.

Identification of zinc finger MIZ-type containing 2 as an oncoprotein enhancing NAD-dependent protein deacetylase sirtuin-1 deacetylase activity to regulate Wnt and Hippo pathways in non-small-cell lung cancer

BackgroundZinc finger MIZ-type containing 2 (ZMIZ2) can function as a coactivator and participate in the progression of certain malignant tumors; however, its expression and underlying molecular mechanism in non-small-cell lung cancer (NSCLC) remains unknown. In this study, we aim to analyze the expression of ZMIZ2 and its tumorigenic function in NSCLC, identifying its related factors.MethodsZMIZ2 expression in NSCLC tissue samples and cell lines was examined using immunohistochemistry and western blotting; its biological role was investigated using in vivo and in vitro assays. The association between ZMIZ2 and NAD-dependent protein deacetylase sirtuin-1 (SIRT1) was demonstrated using mass spectrometry and immunoprecipitation experiments. Kyoto Encyclopedia of Genes and Genomes Pathway (KEGG)-based enrichment analysis, luciferase reporter assay, and real-time quantitative polymerase chain reaction (RT–qPCR) were conducted to verify the impact of ZMIZ2–SIRT1 combination on Hippo/Wnt pathways.ResultsZMIZ2 was highly expressed in NSCLC and positively associated with advanced pTNM staging, lymph node metastasis, and poor overall survival. Functional experiments revealed that ZMIZ2 promotes the proliferation, migration, and invasiveness of lung cancer cells—establishing its role as a promoter of oncogenes. Molecular mechanism studies identified SIRT1 as an assisted key factor interacting with ZMIZ2. KEGG enrichment analysis revealed that ZMIZ2 is closely related to Wnt/Hippo pathways; ZMIZ2–SIRT1 interaction enhanced SIRT1 deacetylase activity. Direct downregulation of intranuclear β-catenin and yes-associated protein (YAP) acetylation levels occurred independently of upstream proteins in Wnt/Hippo pathways; transcriptional activities of β-catenin-transcription factor 4 (TCF4) and YAP–TEA domain family transcription factors (TEADs) were amplified.ConclusionsZMIZ2 promotes the malignant phenotype of lung cancer by regulating Wnt/Hippo pathways through SIRT1, providing an experimental basis for discovering novel biomarkers and developing tumor-targeted drugs.Graphical

Enthalpy of the Cerium (Ce) Chemi-Ionization Reaction and CeO+, CeC+, and CeCO+ Bond Energies Determined by Energy-Resolved Guided Ion Beam Mass Spectrometry Experiments.

Cerium (Ce) oxide, carbide, and carbonyl cation bond energies and the exothermicity of the Ce chemi-ionization (CI) reaction with atomic oxygen were investigated using guided-ion beam tandem mass spectrometry (GIBMS). The kinetic energy dependent product cross sections for reactions of Ce+ with O2, CO, and CO2 and CeO+ with O2, CO, Xe, and Ar were measured using GIBMS. For the reactions of Ce+ with O2 and CO2, CeO+ is formed through an exothermic reaction, whereas CeO+ formation is endothermic in the reaction with CO. This reaction also forms CeC+ and CeCO+ is formed in the reaction of Ce+ with CO2, both in endothermic processes. Reactions of CeO+ with all four gases led to endothermic collision-induced dissociation as well as exchange reactions to form CeO2+ for the O2 and CO reactants. Bond dissociation energies (BDEs) at 0 K were determined through analyses of the kinetic energy dependent cross sections for all endothermic reactions. We determined that the BDEs of CeO+, CeC+, and CeCO+ are 8.46 ± 0.15, 3.93 ± 0.16, and ≥0.25 ± 0.07 eV, respectively, where the CeO+ BDE is a weighted average of 6 independent 0 K threshold measurements. The CeO+ BDE is combined with the ionization energy (IE) of Ce to determine an exothermicity of 2.91 ± 0.15 eV for the Ce + O → CeO+ + e- chemi-ionization reaction. Combined with neutral BDEs from the literature, the thermochemistry determined here also provides IE(CeO) = 5.03 ± 0.12 eV and IE(CeC) = 6.50 ± 0.16 eV. Theoretical calculations for ground and some excited states were performed for CeO+, CeC+, and CeCO+ to provide insight into the bonding and a comparison with the experimentally determined BDEs for each molecule.

DUS4L suppresses invasion and metastasis in LUAD via modulation of PI3K/AKT and ERK/MAPK signaling through GRB2

ObjectiveLimited research has focused on the role of dihydrouridine synthases (DUS) family members in human tumors. Our previous findings indicated an impact of dihydrouridine synthase 4 like (DUS4L) on cell proliferation and apoptosis in lung adenocarcinoma (LUAD) A549 cell, yet its broader functions and regulatory mechanisms in LUAD remain elusive. MethodsUsing a LUAD tissue microarray and immunohistochemical (IHC) staining, we validated variations in DUS4L protein expression levels among LUAD patients and assessed its clinical significance. Additional experiments using short hairpin RNA (shRNA) against DUS4L (sh-DUS4L-2), LUAD cell lines, cell function assays (including wound healing, transwell migration and invasion, colony formation, and apoptosis assays), and mouse tumor xenografts were performed to examine the biological roles of DUS4L in LUAD progression. RNA sequencing, proteomic analyses, mass spectrometry, and co-immunoprecipitation experiments were conducted to identify and validate DUS4L-regulated downstream target genes and signaling pathways. ResultsWe identified a consistent upregulation of DUS4L in LUAD tissues. In vitro and in vivo experiments underscored the inhibitory effect of DUS4L downregulation on LUAD progression, including migration, invasion, and proliferation. Mechanistically, DUS4L was found to interact with the signaling molecule GRB2, promoting LUAD progression and metastasis by inducing epithelial-mesenchymal transition (EMT) via the PI3K/AKT and ERK/MAPK pathways. ConclusionOur results establish the functional role of DUS4L in driving the progression and metastasis of LUAD, implicating its potential as a candidate therapeutic target for LUAD.

Chromophore Optimization in Organometallic Au(III) Cys Arylation of Peptides and Proteins for 266 nm Photoactivation.

Cysteine is the most reactive naturally occurring amino acid due to the presence of a free thiol, presenting a tantalizing handle for covalent modification of peptides/proteins. Although many mass spectrometry experiments could benefit from site-specific modification of Cys, the utility of direct arylation has not been thoroughly explored. Recently, Spokoyny and co-workers reported a Au(III) organometallic reagent that robustly arylates Cys and tolerates a wide variety of solvents and conditions. Given the chromophoric nature of aryl groups and the known susceptibility of carbon-sulfur bonds to photodissociation, we set out to identify an aryl group that could efficiently cleave Cys carbon-sulfur bonds at 266 nm. A streamlined workflow was developed to facilitate rapid examination of a large number of aryls with minimal sample using a simple test peptide, RAAACGVLK. We were able to identify several aryl groups that yield abundant homolytic photodissociation of the adjacent Cys carbon-sulfur bonds with short activation times (<10 ms). In addition, we characterized the radical products created by photodissociation by subjecting the product ions to further collisional activation. Finally, we tested Cys arylation with human hemoglobin, identified reaction conditions that facilitate efficient modification of intact proteins, and evaluated the photochemistry and activation of these large radical ions.

A Mass Spectrometry Experiment on the Degrees of Freedom Effect

A Mass Spectrometry Experiment on the Degrees of Freedom Effect

Ultrathin ZrO2 thickness control on TiO2/ZrO2 core/shell nanoparticles using ZrO2 atomic layer deposition and etching

Atomic layer deposition (ALD) and atomic layer etching (ALE) techniques were used to control the ZrO2 shell thickness on TiO2/ZrO2 core/shell nanoparticles. ALD and ALE were performed at 200 °C while the nanoparticles were agitated using a rotary reactor. To increase the ZrO2 shell thickness, ZrO2 ALD films were deposited using sequential exposures of tetrakis(dimethylamino) zirconium and H2O. Ex situ analysis using transmission electron microscopy (TEM) observed the growth of the ZrO2 shells. The ZrO2 ALD led to more spherical ZrO2 shells on the crystalline and irregular TiO2 cores. The ZrO2 ALD on the nanoparticles had a growth rate of 0.9 ± 0.1 Å/cycle. Tunable ZrO2 coatings were observed with thicknesses ranging from 5.9 to 27.1 nm after 240 ZrO2 ALD cycles. To demonstrate the decrease in the ZrO2 shell thickness, the ZrO2 film was then etched using sequential hydrogen fluoride (HF) and TiCl4 exposures. Quadrupole mass spectrometry experiments performed in a separate reactor identified the volatile products during ZrO2 ALE. H2O was monitored during HF exposures, and ZrCl4 etch products and TiFxCly ligand-exchange products were observed during TiCl4 exposures. Ex situ TEM studies revealed that the ZrO2 shells remained spherical during ZrO2 ALE. The ZrO2 ALE on the nanoparticles had an etch rate of 6.5 ± 0.2 Å/cycle. Tunable ZrO2 coatings were produced from 27.1 down to 7.6 nm using 30 ZrO2 ALE cycles. This study demonstrated that ZrO2 ALD and ZrO2 ALE can control the thickness of ZrO2 shells on TiO2/ZrO2 core/shell nanoparticles without inducing nanoparticle agglomeration.