Tools for investigating O-GlcNAc in signaling and other fundamental biological pathways.

Cells continuously fine-tune signaling pathway proteins to match nutrient and stress levels in their local environment by modifying intracellular proteins with O-linked N-acetylglucosamine (O-GlcNAc) sugars, an essential process for cell survival and growth. The small size of these monosaccharide modifications poses a challenge for functional determination, but the chemistry and biology communities have together created a collection of precision tools to study these dynamic sugars. This review presents the major themes by which O-GlcNAc influences signaling pathway proteins, including G-protein coupled receptors, growth factor signaling, mitogen-activated protein kinase (MAPK) pathways, lipid sensing, and cytokine signaling pathways. Along the way, we describe in detail key chemical biology tools that have been developed and applied to determine specific O-GlcNAc roles in these pathways. These tools include metabolic labeling, O-GlcNAc-enhancing RNA aptamers, fluorescent biosensors, proximity labeling tools, nanobody targeting tools, O-GlcNAc cycling inhibitors, light-activated systems, chemoenzymatic labeling, and nutrient reporter assays. An emergent feature of this signaling pathway meta-analysis is the intricate interplay between O-GlcNAc modifications across different signaling systems, underscoring the importance of O-GlcNAc in regulating cellular processes. We highlight the significance of O-GlcNAc in signaling and the role of chemical and biochemical tools in unraveling distinct glycobiological regulatory mechanisms. Collectively, our field has determined effective strategies to probe O-GlcNAc roles in biology. At the same time, this survey of what we do not yet know presents a clear roadmap for the field to use these powerful chemical tools to explore cross-pathway O-GlcNAc interactions in signaling and other major biological pathways.

Fluorescent Janus emulsions for biosensing of Listeria monocytogenes.

Here we report a sensing method for Listeria monocytogenes based on the agglutination of all-liquid Janus emulsions. This two-dye assay enables the rapid detection of trace Listeria in less than 2 h via an emissive signal produced in response to Listeria binding. The biorecognition interface between the Janus emulsions is assembled by attaching antibodies to a functional surfactant polymer with a tetrazine/transcyclooctene click reaction. The strong binding between Listeria and the Listeria antibody located at the hydrocarbon surface of the emulsions results in the tilting of the Janus structure from its equilibrium position to produce emission that would ordinarily be obscured by a blocking dye. This method provides rapid and inexpensive Listeria detection with high sensitivity (<100 CFU/mL in 2 h) that can be paired with many antibody or related recognition elements to create a new class of biosensors.

ABCs of Faraday Rotation in Organic Materials.

Faraday rotation is a magneto-optical effect central to a number of commercial technologies including optical isolation and magneto-optical imaging. Today, the performance needs of these technologies are met by inorganic materials containing paramagnetic heavy elements. However, organic thin films are increasingly being evaluated as replacement materials, promising higher magneto-optical performance and facile fabrication of structures that enable expanded applications. Despite being an object of research for more than 175 years, our understanding of the Faraday effect in solid-state organic materials remains incomplete, hindering our attempts to methodically improve magneto-optical performance. This Perspective aims to place several recent advances in the field of thin-film organic Faraday rotators within the well-established theoretical framework developed by solution-state magnetic circular dichroism spectroscopists: the Faraday A, B, and C terms. Through careful consideration of these quantum mechanical mechanisms in example molecules, an intuitive understanding of the impact of chemical structure in thin-film Faraday rotators can be achieved, including the critical roles of molecular symmetry, rigidity, absorptivity, and magnetism. Future work seeking to maximize the magneto-optical performance of organic thin films may more readily evaluate candidate chromophores based on the Faraday A, B, and C term framework presented herein.

Molecular Platform for Fast Low-Voltage Nanoelectromechanical Switching.

The use of molecules as active components to build nanometer-scale devices inspires emerging device concepts that employ the intrinsic functionality of molecules to address longstanding challenges facing nanoelectronics. Using molecules as controllable-length nanosprings, here we report the design and operation of a nanoelectromechanical (NEM) switch which overcomes the typical challenges of high actuation voltages and slow switching speeds for previous NEM technologies. Our NEM switches are hierarchically assembled using a molecular spacer layer sandwiched between atomically smooth electrodes, which defines a nanometer-scale electrode gap and can be electrostatically compressed to repeatedly modulate the tunneling current. The molecular layer and the top electrode structure serve as two degrees of design freedom with which to independently tailor static and dynamic device characteristics, enabling simultaneous low turn-on voltages (sub-3 V) and short switching delays (2 ns). This molecular platform with inherent nanoscale modularity provides a versatile strategy for engineering diverse high-performance and energy-efficient electromechanical devices.

Large Faraday Rotation in Optical-Quality Phthalocyanine and Porphyrin Thin Films.

The magneto-optical phenomenon known as Faraday rotation involves the rotation of plane-polarized light as it passes through an optical medium in the presence of an external magnetic field oriented parallel to the direction of light propagation. Faraday rotators find applications in optical isolators and magnetic-field imaging technologies. In recent years, organic thin films comprised of polymeric and small-molecule chromophores have demonstrated Verdet constants, which measure the magnitude of rotation at a given magnetic field strength and material thickness, that exceed those found in conventional inorganic crystals. We report herein the thin-film magnetic circular birefringence (MCB) spectra and maximum Verdet constants of several commercially available and newly synthesized phthalocyanine and porphyrin derivatives. Five of these species achieved maximum Verdet constant magnitudes greater than 105 deg T-1 m-1 at wavelengths between 530 and 800 nm. Notably, a newly reported zinc(II) phthalocyanine derivative (ZnPc-OT) reached a Verdet constant of -33 × 104 deg T-1 m-1 at 800 nm, which is among the largest reported for an organic material, especially for an optical-quality thin film. The MCB spectra are consistent with resonance-enhanced Faraday rotation in the region of the Q-band electronic transition common to porphyrin and phthalocyanine derivatives, and the Faraday A-term describes the electronic origin of the magneto-optical activity. Overall, we demonstrate that phthalocyanines and porphyrins are a class of rationally designed magneto-optical materials suitable for applications demanding large Verdet constants and high optical quality.

GlycoID Proximity Labeling to Identify O-GlcNAcylated Protein Interactomes in Live Cells.

Cells continuously remodel their intracellular proteins with the monosaccharide O-linked N-acetylglucosamine (O-GlcNAc) to regulate metabolism, signaling, and stress. This protocol describes the use of GlycoID tools to capture O-GlcNAc dynamics in live cells. GlycoID constructs contain an O-GlcNAc binding domain linked to a proximity labeling domain and a subcellular localization sequence. When expressed in mammalian cells, GlycoID tracks changes in O-GlcNAc-modified proteins and their interactomes in response to chemical induction with biotin over time. Pairing the subcellular localization of GlycoID with the chemical induction of activity enables spatiotemporal studies of O-GlcNAc biology during cellular events such as insulin signaling. However, optimizing intracellular labeling experiments requires attention to several variables. Here, we describe two protocols to adapt GlycoID methods to a cell line and biological process of interest. Next, we describe how to conduct a semiquantitative proteomic analysis of O-GlcNAcylated proteins and their interactomes using insulin versus glucagon signaling as a sample application. This articles aims to establish baseline GlycoID protocols for new users and set the stage for widespread use over diverse cellular applications for the functional study of O-GlcNAc glycobiology. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Expression of targeted GlycoID constructs to verify subcellular location and labeling activity in mammalian cells Basic Protocol 2: GlycoID labeling in live HeLa cells for O-GlcNAc proteomic comparisons.

Pseudo-fivefold diffraction symmetries in tetrahedral packing.

We review the way in which atomic tetrahedra composed of metallic elements pack naturally into fused icosahedra. Orthorhombic, hexagonal, and cubic intermetallic crystals based on this packing are all shown to be united in having pseudo-fivefold rotational diffraction symmetry. A unified geometric model involving the 600-cell is presented: the model accounts for the observed pseudo-fivefold symmetries among the different Bravais lattice types. The model accounts for vertex-, edge-, polygon-, and cell-centered fused-icosahedral clusters. Vertex-centered and edge-centered types correspond to the well-known pseudo-fivefold symmetries in Ih and D5h quasicrystalline approximants. The concept of a tetrahedrally-packed reciprocal space cluster is introduced, the vectors between sites in this cluster corresponding to the principal diffraction peaks of fused-icosahedrally-packed crystals. This reciprocal-space cluster is a direct result of the pseudosymmetry and, just as the real-space clusters, can be rationalized by the 600-cell. The reciprocal space cluster provides insights for the Jones model of metal stability. For tetrahedrally-packed crystals, Jones zone faces prove to be pseudosymmetric with one another. Lower and upper electron per atom bounds calculated for this pseudosymmetry-based Jones model are shown to accord with the observed electron counts for a variety of Group 10-12 tetrahedrally-packed structures, among which are the four known Cu/Cd intermetallic compounds: CdCu2, Cd3Cu4, Cu5Cd8, and Cu3Cd10. The rationale behind the Jones lower and upper bounds is reviewed. The crystal structure of Zn11Au15Cd23, an example of a 1:1 MacKay cubic quasicrystalline approximant based solely on Groups 10-12 elements is presented. This compound crystallizes in Im3 (space group no. 204) with a = 13.842(2) Å. The structure was solved with R1 = 3.53 %, I > 2σ; = 5.33 %, all data with 1282/0/38 data/restraints/parameters.

Exploring the Complementarity of Measures of Instructional Practices.

The assessment of instructional quality has been and continues to be a desirable, yet difficult endeavor in higher education. The development of new teaching evaluation frameworks along with instruments to measure various aspects of teaching practices holds promise. The challenge rests in the implementation of these frameworks and measures in authentic settings. Part of this challenge is for instructors, researchers, and administrators to parse through and select a meaningful set of tools from the plethora of existing instruments. In this study, we aim to start clarifying the landscape of measures of instructional practice by exploring the complementarity of two existing instruments: the Classroom Observation Protocol for Undergraduate STEM (COPUS) and the Learner-Centered Teaching Rubrics (LCTR). We collected classroom observations and course artifacts from 28 science instructors from research-intensive institutions across the United States. Results show the need to use both instruments to capture nuanced and comprehensive description of a faculty member's instructional practice. This study highlights the messiness of measuring instructional quality and the need to explore the implementation of teaching evaluation frameworks and measures of instructional practices in authentic settings.

Spatiotemporal Proximity Labeling Tools to Track GlcNAc Sugar-Modified Functional Protein Hubs during Cellular Signaling.

A fundamental mechanism that all eukaryotic cells use to adapt to their environment is dynamic protein modification with monosaccharide sugars. In humans, O-linked N-acetylglucosamine (O-GlcNAc) is rapidly added to and removed from diverse protein sites as a response to fluctuating nutrient levels, stressors, and signaling cues. Two aspects remain challenging for tracking functional O-GlcNAc events with chemical strategies: spatial control over subcellular locations and time control during labeling. The objective of this study was to create intracellular proximity labeling tools to identify functional changes in O-GlcNAc patterns with spatiotemporal control. We developed a labeling strategy based on the TurboID proximity labeling system for rapid protein biotin conjugation directed to O-GlcNAc protein modifications inside cells, a set of tools called "GlycoID." Localized variants to the nucleus and cytosol, nuc-GlycoID and cyt-GlycoID, labeled O-GlcNAc proteins and their interactomes in subcellular space. Labeling during insulin and serum stimulation revealed functional changes in O-GlcNAc proteins as soon as 30 min following signal initiation. We demonstrated using proteomic analysis that the GlycoID strategy captured O-GlcNAcylated "activity hubs" consisting of O-GlcNAc proteins and their associated protein-protein interactions. The ability to follow changes in O-GlcNAc hubs during physiological events such as insulin signaling allows these tools to determine the mechanisms of glycobiological cell regulation. Our functional O-GlcNAc data sets in human cells will be a valuable resource for O-GlcNAc-driven mechanisms.

C-Term Faraday Rotation in Metallocene Containing Thin Films.

The Faraday effect is a magneto-optical (MO) phenomenon that causes the plane of linearly polarized light to rotate when passing through a medium subjected to a parallel magnetic field. Informed by the established quantum mechanical model developed by Buckingham and Stephens, we sought to identify molecules that would exhibit large MO responses. Magnetic circular dichroism studies of ferrocenium in the 1970s revealed its potential as an MO material; however, it has not been evaluated in the context of Faraday rotation and thin-film optical applications. Herein, we report near-infrared (NIR) Faraday rotation in thin films of decamethylferrocenium/poly(methyl methacrylate) composites with maximum Verdet constants of -3.45 × 104 deg T-1 m-1 at 810 nm (absorbance = 0.09) and -1.44 × 104 deg T-1 m-1 at 870 nm (absorbance = 0.01). These polymer-metallocene thin films deliver larger Verdet constants than commercially used NIR inorganic Faraday rotators and are facile and inexpensive to produce. The temperature dependence and distinct lineshape of the MO responses observed in decamethylferrocenium radical cations, decamethylmanganocene, and chromocene are in accordance with the quantum mechanical model. The observation of a strong C-term Faraday rotation in solid-state organometallic materials provides the groundwork for the development of high-performance metallocene-based Faraday rotators.

Effectiveness and Feasibility of Pharmacist-Driven Penicillin Allergy De-Labeling Pilot Program without Skin Testing or Oral Challenges.

Documented penicillin allergies have been associated with an increased risk of adverse outcomes. The goal of this project was to assess the effectiveness and feasibility of a pharmacist-led penicillin allergy "de-labeling" process that does not involve labor-intensive skin testing or direct oral challenges. Adult patients with penicillin allergies were identified and interviewed by an infectious diseases pharmacy resident during a 3-month pilot period. Using an evidence-based standardized checklist, the pharmacist determined if an allergy qualified for de-labeling. In total, 66 patients were interviewed during the pilot period. The average time spent was 5.2 min per patient interviewed. Twelve patients (18%) met the criteria for de-labeling and consented to the removal of the allergy. Four patients (6%) met the criteria but declined removal of the allergy. In brief, 58.3% of patients (7/12) who were de-labeled and 50% of patients (2/4) who declined de-labeling but had their allergy updated to reflect intolerance were subsequently prescribed beta-lactam antibiotics and all (9/9, 100%) were able to tolerate these agents. A pharmacist-led penicillin allergy de-labeling process utilizing a standardized checklist is an effective and feasible method for removing penicillin allergies in patients without a true allergy.

Janus Emulsion Biosensors for Anti-SARS-CoV-2 Spike Antibody.

The spread of the COVID-19 pandemic around the world has revealed that it is urgently important to develop rapid and inexpensive assays for antibodies in general and anti-SARS-CoV-2 IgG antibody (anti-SARS-CoV-2 spike glycoprotein S1 antibody) in particular. Herein we report a method to detect the anti-SARS-CoV-2 spike antibody level by using Janus emulsions or Janus particles as biosensors. Janus emulsions are composed of two immiscible hydrocarbon and fluorocarbon oils. The hydrocarbon/water interfaces are functionalized with a secondary antibody of IgG protein and SARS-CoV-2 spike receptor binding domain (RBD), to produce two different Janus emulsions. Mixtures of these Janus droplets enable the detection of the anti-SARS-CoV-2 spike IgG antibody in an agglutination assay caused by the antibody's binding to both the secondary antibody of IgG antibody and SARS-CoV-2 spike protein RBD. Both qualitative optical images and quantitative fluorescence spectra are able to detect the level of anti-SARS-CoV-2 spike antibody at concentrations as low as 0.2 µg/mL in 2 h. The detection results of clinical human serum samples using this agglutination assay confirm that this method is applicable to clinical samples with good sensitivity and specificity. The reported method is generalizable and can be used to detect other analytes by attaching different biomolecular recognition elements to the surface of the Janus droplets.

Chemical Structure of Fe-Ni Nanoparticles for Efficient Oxygen Evolution Reaction Electrocatalysis.

Bimetallic iron-nickel-based nanocatalysts are perhaps the most active for the oxygen evolution reaction (OER) in alkaline electrolytes. Recent developments in literature have suggested that the ratio of iron and nickel in Fe-Ni thin films plays an essential role in the performance and stability of the catalysts. In this work, the metallic ratio of iron to nickel was tested in alloy bimetallic nanoparticles. Similar to thin films, nanoparticles with iron-nickel atomic compositions where the atomic iron percentage is ≤50% outperformed nanoparticles with iron-nickel ratios of >50%. Nanoparticles of Fe20Ni80, Fe50Ni50, and Fe80Ni20 compositions were evaluated and demonstrated to have overpotentials of 313, 327,, and 364 mV, respectively, at a current density of 10 mA/cm2. While the Fe20Ni80 composition might be considered to have the best OER performance at low current densities, Fe50Ni50 was found to have the best current density performance at higher current densities, making this composition particularly relevant for electrolysis conditions. However, when stability was evaluated through chronoamperometry and chronopotentiometry, the Fe80Ni20 composition resulted in the lowest degradation rates of 2.9 µA/h and 17.2 µV/h, respectively. These results suggest that nanoparticles with higher iron and lower nickel content, such as the Fe80Ni20 composition, should be still taken into consideration while optimizing these bimetallic OER catalysts for overall electrocatalytic performance. Characterization by electron microscopy, diffraction, and X-ray spectroscopy provides detailed chemical and structural information on as-synthesized nanoparticle materials.

What Can a Urine Drug Screening Immunoassay Really Tell Us?

Urine drug screening has become standard of care in many medical practice settings to assess compliance, detect misuse, and/or to provide basis for medical or legal action. The antibody-based enzymatic immunoassays used for qualitative analysis of urine have significant drawbacks that clinicians are often not aware of. Recent literature suggests that there is a lack of understanding of the shortcomings of these assays by clinicians who are ordering and/or interpreting them. This article addresses the state of each of the individual immunoassays that are most commonly used today in order to help the reader become proficient in the interpretation and application of the results. Some literature already exists regarding sources of "false positives" and "false negatives," but none seem to present the material with the practicing clinician in mind. This review aims to avoid overwhelming the reader with structures and analytical chemistry. The reader will be presented relevant clinical knowledge that will facilitate appropriate interpretation of immunoassays regardless of practice settings. Using this review as a learning tool and a reference, clinicians will be able to interpret the results of commonly used immunoassays in an evidence-based, informed manner and minimize the negative impact that misinterpretation has on patient care.

Relationships between five factor personality variables, workplace accidents, and self-efficacy.

Past research has examined the relationships between personality variables and workplace accidents; however, few studies have examined these relationships using the Five Factor Model of personality and trait-state relationships have not been examined in this context. The present study was undertaken to examine the relationships between personality characteristics based on the Five Factor Model, workplace accidents, and self-efficacy. To examine these relationships, personality, workplace accident, and self-efficacy data were collected from 202 undergraduate volunteers (134 women and 68 men) at a large midwestern urban university. The mean age of participants was 20.9 yr. (SD=5.0). The results indicated that scores on Agreeableness and Conscientiousness were significantly correlated with workplace accidents as were the self-efficacy variables. Neuroticism and Agreeableness were significantly related to self-efficacy.

Carbonylative enantioselective meso-desymmetrization of cis-epoxides to trans-ß-lactones: effect of salen-ligand electronic variation on enantioselectivity.

Carbonylation catalysts for the desymmetrization of meso-epoxides yielding enantioenriched trans-ß-lactones are reported. Fine-tuning the electronic properties of the ligand further improved enantioselectivity. The sterics of the substrate dictated whether a given electronic variation decreased or increased enantioenrichment, revealing an unexpected relationship between the substrate's steric environment and the electronic nature of the optimal catalyst.