Candidate Tear-Based Uveitis Biomarkers in Children with JIA Based on Arthritis Activity and Topical Corticosteroid Use.

Background: Uveitis is an inflammatory ocular disease secondary to disruption of the retinal pigmented epithelium (RPE) and blood retinal barrier (BRB). Known clinical factors do not accurately predict uveitis risk in Juvenile Idiopathic Arthritis (JIA). Tear fluid is easily obtained for biomarker study.

Mechanistic insight into benzylidene-directed glycosylation reactions using cryogenic infrared spectroscopy.

The stereoselective formation of 1,2- glycosidic linkages is challenging. The currently most widely used strategy for their installation uses 4,6--benzylidene-protected building blocks. The stereoselectivity of this reaction is thought to be driven by a covalent intermediate, which reacts via an S2 mechanism.

Direct Experimental Characterization of a Sialyl Cation.

Sialic acids are monosaccharide residues involved in several biological processes. Controlling the stereoselectivity of sialylation reactions is challenging and mechanistic studies on the structure of its intermediate, the sialyl cation, are scarce. Here it is shown that a sialyl cation can be generated and isolated from an ionized sialic acid precursor.

The impact of side-chain fluorination on proton-bound phenylalanine dimers: a cryogenic infrared spectroscopic study.

The incorporation of fluorine into amino acids is an important strategy to produce tailored building blocks with unique properties for peptide-based materials. Phenylalanine is frequently modified due to its role in cation-π interactions and the formation of amyloid fibres. Previous studies have utilized gas-phase vibrational spectroscopy to study interactions between canonical amino acids.

Evolutionary origins and innovations sculpting the mammalian PRPS enzyme complex.

The phosphoribosyl pyrophosphate synthetase (PRPS) enzyme conducts a chokepoint reaction connecting central carbon metabolism and nucleotide production pathways, making it essential for life. Here, we show that the presence of multiple PRPS-encoding genes is a hallmark trait of eukaryotes, and we trace the evolutionary origins and define the individual functions of each of the five mammalian PRPS homologs - three isozymes (one testis-restricted) and two non-enzymatic associated proteins (APs) - which we demonstrate operate together as a large molecular weight complex capable of attaining a heterogeneous array of functional multimeric configurations. Employing a repertoire of isogenic fibroblast clones in all viable individual or combinatorial assembly states, we define preferential interactions between subunits, and we show that cells lacking PRPS2, PRPSAP1, and PRPSAP2 render PRPS1 into aberrant homo-oligomeric assemblies with diminished metabolic flux and impaired proliferative capacity.

Structure and Conformation Determine Gas-Phase Infrared Spectra of Detergents.

Native mass spectrometry of membrane proteins relies on non-ionic detergents which protect the protein during transfer from solution into the gas phase. Once in the gas phase, the detergent micelle must be efficiently removed, which is usually achieved by collision-induced dissociation (CID). Recently, infrared multiple photon dissociation (IRMPD) has emerged as an alternative activation method for the analysis of membrane proteins, which has led to a growing interest in detergents that efficiently absorb infrared light.

Dysregulated innate immune signaling cooperates with RUNX1 mutations to transform an MDS-like disease to AML.

Dysregulated innate immune signaling is linked to preleukemic conditions and myeloid malignancies. However, it is unknown whether sustained innate immune signaling contributes to malignant transformation. Here we show that cell-intrinsic innate immune signaling driven by miR-146a deletion (miR-146a), a commonly deleted gene in myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML), cooperates with mutant RUNX1 (RUNX1) to initially induce marrow failure and features of MDS.

Infrared Multiphoton Dissociation Enables Top-Down Characterization of Membrane Protein Complexes and G Protein-Coupled Receptors.

Membrane proteins are challenging to analyze by native mass spectrometry (MS) as their hydrophobic nature typically requires stabilization in detergent micelles that are removed prior to analysis via collisional activation. There is however a practical limit to the amount of energy which can be applied, which often precludes subsequent characterization by top-down MS. To overcome this barrier, we have applied a modified Orbitrap Eclipse Tribrid mass spectrometer coupled to an infrared laser within a high-pressure linear ion trap.

Hydrogen Tag Shifts as Vibrational Reporters for Positional Isomers of Formylphenides: Surprising Mobility of the Carbanion Center upon Collisional Decarboxylation of the Parent Benzoates.

Infrared photodissociation of weakly bound "mass tags" is widely used to determine the structures of ions by analyzing their vibrational spectra. Molecular hydrogen is a common choice for tagging in cryogenic radio-frequency ion traps. Although the H molecules can introduce distortions in the target species, we demonstrate an advantage of H tagging in the analysis of positional isomers adopted by the molecular anions derived from decarboxylation of formylbenzoates.

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.

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.

Prenatal SARS-CoV-2 infection alters postpartum human milk-derived extracellular vesicles.

Human milk-derived extracellular vesicles (HMEVs) are crucial functional components in breast milk, contributing to infant health and development. Maternal conditions could affect HMEV cargos; however, the impact of SARS-CoV-2 infection on HMEVs remains unknown. This study evaluated the influence of SARS-CoV-2 infection during pregnancy on postpartum HMEV molecules.

Infrared Multiphoton Dissociation Enables Top-Down Characterization of Membrane Protein Complexes and G Protein-Coupled Receptors.

Membrane proteins are challenging to analyze by native mass spectrometry (MS) as their hydrophobic nature typically requires stabilization in detergent micelles that are removed prior to analysis via collisional activation. There is however a practical limit to the amount of energy which can be applied, which often precludes subsequent characterization by top-down MS. To overcome this barrier, we have applied a modified Orbitrap Eclipse Tribrid mass spectrometer coupled to an infrared laser within a high-pressure linear ion trap.

Repression of TRIM13 by chromatin assembly factor CHAF1B is critical for AML development.

Acute myeloid leukemia (AML) is an aggressive blood cancer that stems from the rapid expansion of immature leukemic blasts in the bone marrow. Mutations in epigenetic factors represent the largest category of genetic drivers of AML. The chromatin assembly factor CHAF1B is a master epigenetic regulator of transcription associated with self-renewal and the undifferentiated state of AML blasts.

Paralog-specific signaling by IRAK1/4 maintains MyD88-independent functions in MDS/AML.

Dysregulation of innate immune signaling is a hallmark of hematologic malignancies. Recent therapeutic efforts to subvert aberrant innate immune signaling in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) have focused on the kinase IRAK4. IRAK4 inhibitors have achieved promising, though moderate, responses in preclinical studies and clinical trials for MDS and AML.

Characterization of Oxidation Products from HOCl Uptake by Microhydrated Methionine Anions Using Cryogenic Ion Vibrational Spectroscopy.

The oxidation of the amino acid methionine (Met) by hypochlorous acid (HOCl) to yield methionine sulfoxide (MetO) has been implicated in both the interfacial chemistry of tropospheric sea spray aerosols and the destruction of pathogens in the immune system. Here, we investigate the reaction of deprotonated methionine water clusters, Met·(HO), with HOCl and characterize the resulting products using cryogenic ion vibrational spectroscopy and electronic structure calculations. Capture of the MetO oxidation product in the gas phase requires the presence of water molecules attached to the reactant anion.

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.

Decoding the Fucose Migration Product during Mass-Spectrometric analysis of Blood Group Epitopes.

Fucose is a signaling carbohydrate that is attached at the end of glycan processing. It is involved in a range of processes, such as the selectin-dependent leukocyte adhesion or pathogen-receptor interactions. Mass-spectrometric techniques, which are commonly used to determine the structure of glycans, frequently show fucose-containing chimeric fragments that obfuscate the analysis.

Establishing carbon-carbon double bond position and configuration in unsaturated fatty acids by gas-phase infrared spectroscopy.

Fatty acids are an abundant class of lipids that are characterised by wide structural variation including isomeric diversity arising from the position and configuration of functional groups. Traditional approaches to fatty acid characterisation have combined chromatography and mass spectrometry for a description of the composition of individual fatty acids while infrared (IR) spectroscopy has provided insights into the functional groups and bond configurations at the bulk level. Here we exploit universal 3-pyridylcarbinol ester derivatization of fatty acids to acquire IR spectra of individual lipids as mass-selected gas-phase ions.

Insights into pulmonary phosphate homeostasis and osteoclastogenesis emerge from the study of pulmonary alveolar microlithiasis.

Pulmonary alveolar microlithiasis is an autosomal recessive lung disease caused by a deficiency in the pulmonary epithelial Npt2b sodium-phosphate co-transporter that results in accumulation of phosphate and formation of hydroxyapatite microliths in the alveolar space. The single cell transcriptomic analysis of a pulmonary alveolar microlithiasis lung explant showing a robust osteoclast gene signature in alveolar monocytes and the finding that calcium phosphate microliths contain a rich protein and lipid matrix that includes bone resorbing osteoclast enzymes and other proteins suggested a role for osteoclast-like cells in the host response to microliths. While investigating the mechanisms of microlith clearance, we found that Npt2b modulates pulmonary phosphate homeostasis through effects on alternative phosphate transporter activity and alveolar osteoprotegerin, and that microliths induce osteoclast formation and activation in a receptor activator of nuclear factor-κB ligand and dietary phosphate dependent manner.

The Influence of the Electron Density in Acyl Protecting Groups on the Selectivity of Galactose Formation.

The stereoselective formation of 1,2--glycosidic bonds is a major bottleneck in the synthesis of carbohydrates. We here investigate how the electron density in acyl protecting groups influences the stereoselectivity by fine-tuning the efficiency of remote participation. Electron-rich C4-pivaloylated galactose building blocks show an unprecedented α-selectivity.

Microhydration of the metastable N-protomer of 4-aminobenzoic acid by condensation at 80 K: H/D exchange without conversion to the more stable O-protomer.

4-aminobenzoic acid (4ABA) is a model scaffold for studying solvent-mediated proton transfer. Although protonation at the carboxylic group (O-protomer) is energetically favored in the gas phase, the N-protomer, where the proton remains on the amino group, can be kinetically trapped by electrospray ionization of 4ABA in an aprotic solvent such as acetonitrile. Here, we report the formation of the hydrated deuterium isotopologues of the N-protomers, RND ·(HO), (R = CHCOOD), which are generated by condensing water molecules onto the bare N-protomers in a liquid nitrogen cooled, radiofrequency octopole ion trap at 80 K.