👤 Harry Dym

Resuscitative thoracotomy (RT) is a last-resort intervention for traumatic cardiac arrest or impending cardiovascular collapse. Although outcomes after RT are well described in civilian trauma, data from modern warfare-characterized by high-energy penetrating mechanisms, advanced prehospital care, and rapid evacuation-remain limited. This study evaluated the characteristics and outcomes of RT performed during recent combat operations within a combined military-civilian trauma system. We conducted a retrospective cohort study of all combat casualties who underwent emergency department (ED) RT during the Israel-Hamas conflict (October 27, 2023, to October 27, 2025). Data were extracted from prehospital and ED medical records and postmortem computed tomography reports. RT was defined as a thoracotomy performed in the ED in a pulseless patient with the intent to restore spontaneous circulation. The primary outcome was 30-day survival. Secondary outcomes included return of spontaneous circulation (ROSC) and 24-hour survival. Among 2,335 combat trauma admissions, 27 patients (1.2%) underwent RT. All were young male casualties with penetrating injuries, predominantly from explosive mechanisms (74.1%). Severe trauma was common (ISS ≥25 in 92.6%). Prehospital blood products were administered in 77.8% of patients, and 66.7% arrived at the ED within 60 minutes of injury. ROSC was achieved in 40.7%, of whom 90.9% were transferred to the operating room. Two patients (7.4%) survived to 24 hours and 30 days, both with good neurologic outcomes. No patient who lost pulse before hospital arrival survived. Among modern warfare casualties treated at civilian trauma centers, survival after RT is comparable to that reported in civilian series, despite severe and complex injury patterns. RT should not be considered futile for penetrating abdominal, pelvic, or extremity hemorrhage, even in the presence of associated head injury. In contrast, prehospital circulatory arrest is associated with an extremely poor prognosis.(J Trauma Acute Care Surg. 2026;000:000-000. Copyright © 2026 Wolters Kluwer Health, Inc. All rights reserved.). Show less

The association between autoimmune diseases and type 1 diabetes (T1D) is mostly based on studies among people with T1D at baseline. We assessed the risk of incident T1D among adolescents with other autoimmune diseases. Included were all Israeli adolescents without a history of dysglycemia, aged 16-19 years, undergoing medical evaluation before mandatory military service between January 1996 and December 2016. Data were linked with information on adult-onset T1D from the Israeli National Diabetes Registry. The cohort was dichotomized by the presence of any autoimmune disease. Cox proportional hazards modeling was applied. A total of 1,426,362 people were included, of whom 38,766 (2.7%) had a history of autoimmunity at study entry (10,333 with autoimmune thyroid disease [AITD] and 9,603 with celiac disease). Over 15,810,751 person-years of follow-up, there were 37 and 740 incident cases of T1D among people with and without autoimmunity, respectively, and a crude incident rate of 9.6 and 4.8 cases per 105 person-years, respectively. In a multivariable model adjusted for sex, birth year, and sociodemographic variables, the hazard ratio (HR) for incident T1D among people with autoimmunity was 2.19 (95% CI 1.57-3.04) versus those without. Results persisted when islet autoantibody data were used as mandatory criteria for T1D case definition (HR 2.22, 95% CI 1.13-4.35). The HRs among people with AITD and celiac disease were 3.99 (2.5-6.4) and 2.82 (1.46-5.45), respectively. Autoimmune diseases in late adolescence were associated with an increased risk of T1D in adulthood in both sexes, especially among those with AITD and celiac disease. Show less

Adolescence is a developmental period characterized by heightened plasticity. Yet, how ongoing development affects sensory processing and cognitive function is unclear. We investigated how adolescent (postnatal day 20-42) and adult (postnatal day 60-82) mice differ in performance on a pure tone Go/No-Go auditory discrimination task of varying difficulty. Using dense electrophysiological recordings, we measured spiking activity at single neuron resolution in the auditory cortex while mice were engaged in the task. As compared to adults, adolescent mice showed lower auditory discrimination performance in a difficult task. This difference in performance was due to higher response variability and weaker cognitive control expressed as higher lick bias. Adolescent and adult neuronal responses differed only slightly in representations of pure tones when measured outside the context of learning and the task. However, cortical representations after learning within the context of the task were markedly different. We found differences in stimulus- and choice-related activity at the single neuron level representations, as well as lower population-level decoding of the difficult task in adolescents. Overall, cortical decoding in adolescents was lower and slower, especially for difficult sound discrimination, reflecting immature cortical representations of sounds and choices. Notably, we found age-related differences, which were more pronounced after learning, reflecting the combined impact of age and learning. Our findings highlight distinct neurophysiological and behavioral profiles in adolescence, underscoring the ongoing development of cognitive control mechanisms and cortical plasticity during this sensitive developmental period. Show less

Proper protein targeting to organelles is crucial for maintaining eukaryotic cellular function and homeostasis. This necessity has driven the evolution of specific targeting signals on proteins and the targeting factors that recognize them. A prominent example is peroxisomal matrix proteins, most of which depend on the targeting factor Pex5 to localize and function correctly. Although most Pex5 cargoes contain a peroxisomal targeting signal type 1 (PTS1), they are not all targeted similarly. Some undergo priority targeting, facilitated either by stronger binding to specific subsets of PTS1 signals or by additional interaction interfaces. These observations highlight the extensive complexity of Pex5-mediated targeting. In this study, we reveal that the Saccharomyces cerevisiae (yeast) matrix protein Eci1 can reach peroxisomes and bind Pex5 in the absence of PTS1. By solving the structure of the yeast Pex5-Eci1 complex using cryo-electron microscopy, we identified additional binding interfaces. Our findings provide new insights into the versatile interactions between Pex5 and its cargo, Eci1. More broadly, this work highlights the intricate, dynamic nature of the interactions between cargo factors and their cargoes to meet the complex environment within eukaryotic cells. Show less

Phytoplankton are responsible for half of the global photosynthesis and form vast blooms in aquatic ecosystems. Bloom demise fuels marine microbial life and is suggested to be mediated by programmed cell death (PCD) induced by diverse environmental stressors. Despite its importance, the molecular basis for algal PCD remains elusive. Here, we reveal novel PCD genes conserved across distant algal lineages using cell-to-cell heterogeneity in the response of the diatom Phaeodactylum tricornutum to oxidative stress. Comparative transcriptomics of sorted sensitive and resilient subpopulations following oxidative stress revealed genes directly linked to their contrasting fates of cell death and survival. Comparing these genes with those found in a large-scale mutant screen in the green alga Chlamydomonas reinhardtii identified functionally relevant conserved PCD gene candidates, including the cysteine protease cathepsin X/Z (CPX). CPX mutants in P. tricornutum CPX1 and C. reinhardtii CYSTEINE ENDOPEPTIDASE 12 (CEP12) exhibited resilience to oxidative stress and infochemicals that induce PCD, supporting a conserved function of these genes in algal PCD. Phylogenetic and predictive structural analyses show that CPX is highly conserved in eukaryotes, and algae exhibit strong structural similarity to human Cathepsin X/Z (CTSZ), a protein linked to various diseases. CPX is expressed by diverse algae across the oceans and correlates with upcoming demise events during toxic Pseudo-nitzschia blooms, providing support for its ecological significance. Elucidating PCD components in algae sheds light on the evolutionary origin of PCD in unicellular organisms and on the cellular strategies employed by the population to cope with stressful conditions. Show less

The opioid epidemic continues to grow, placing a significant strain on Emergency Departments (EDs), resulting in patients presenting daily with opioid-related concerns including intoxication, withdrawal, infections, injury, and death. Consequently, in recent years many EDs, including our own, have utilized Emergency Department Observation Units (EDOU) to not only manage withdrawal and overdose, but also initiate long-term treatment. This study aims to evaluate the outcomes of patients with opioid use disorder (OUD) who were managed in our EDOU. This was a retrospective study of patients placed in an EDOU who had the primary diagnosis of OUD in a single large, urban, tertiary academic hospital from May to November 2021. Demographic data and factors related to the ED visit and EDOU actions (e.g., use of peer navigator services, buprenorphine dose and prescription, distribution of naloxone discharge kits, and addiction clinic referral) were analyzed. The primary outcome variables were complications after buprenorphine use (e.g., precipitated withdrawal), the number of repeat ED visits or subsequent hospitalizations within 30 days for both all causes and opioid-related causes, and fatalities within 30 days of EDOU discharge. Twenty-nine patients were identified for chart review. Of these, 59 % were male. The median age was 55 years. Additionally, 93 % of the patients were insured, 66 % had housing, 72 % possessed a phone, and none were employed. During EDOU stays, 48 % [95 % CI 0.2989, 0.6711] of patients received buprenorphine with a total mean dose of 19 mg (SD, 10.6 mg). Upon discharge from the EDOU, 48 % [95 % CI 0.2989, 0.6711] were prescribed buprenorphine, 14 % [95 % CI 0.0451, 0.3257] received a naloxone discharge kit, and 45 % [95 % CI 0.2696, 0.6402] received an addiction clinic appointment. No patients had precipitated withdrawal, serious adverse events, or upgrades to inpatient care. Within 30-days of EDOU discharge, 38 % [95 % CI 0.213, 0.5764] of patients had a subsequent ED visit for any cause, and 6.9 % [95 % CI 1.2, 2.2] had a subsequent hospitalization for any cause. There were no fatalities within 30 days of EDOU discharge. The EDOU can serve as a promising location to provide quality care for patients presenting to the ED with OUD, with minimal adverse effects. There were few subsequent hospitalizations following discharge from the EDOU. Further non-observational studies regarding OUD management in an EDOU setting should be performed to optimize care and improve clinical outcomes and healthcare utilization. Show less

Domains known as von Willebrand factor type D (VWD) are found in extracellular and cell-surface proteins including von Willebrand factor, mucins, and various signaling molecules and receptors. Many VWD domains have a glycine-aspartate-proline-histidine (GDPH) amino-acid sequence motif, which is hydrolytically cleaved post-translationally between the aspartate (Asp) and proline (Pro). The Fc IgG binding protein (FCGBP), found in intestinal mucus secretions and other extracellular environments, contains 13 VWD domains, 11 of which have a GDPH cleavage site. In this study, we investigated the structural and biophysical consequences of Asp-Pro peptide cleavage in a representative FCGBP VWD domain. We found that endogenous Asp-Pro cleavage increases the resistance of the domain to exogenous proteolytic degradation. Tertiary structural interactions made by the newly generated chain termini, as revealed by a crystal structure of an FCGBP segment containing the VWD domain, may explain this observation. Notably, the Gly-Asp peptide bond, upstream of the cleavage site, assumed the cis configuration in the structure. In addition to these local features of the cleavage site, a global organizational difference was seen when comparing the FCGBP segment structure with the numerous other structures containing the same set of domains. Together, these data illuminate the outcome of GDPH cleavage and demonstrate the plasticity of proteins with VWD domains, which may contribute to their evolution for function in a dynamic extracellular environment. Show less

During the development of multimodal pain management protocols, practitioners need to consider the potential risks each treatment modality inherently carries in order to prevent or diminish harmful outcomes. As an example, the part dentists played in the early stages of the opioid epidemic in the United States of America should serve as a cautionary account. By understanding the roots of this crisis, as practitioners we are better equipped to implement the novel analgesic agents available today to optimize post-operative pain control while minimizing any risk of addiction and harm to our communities. It is therefore critical that our colleagues understand the variety of accessible options for pain management to assure that our profession is able to seek adequate and sustainable relief for our post-operative patients. This article will go in depth to explain the analgesic tools practitioners can implement for an effective low-risk protocol, including a combination of NSAIDS and acetaminophen approach, using long-acting local anesthetics such as Exparel, pregabalin, gabapentin, ketamine, dexmedetomidine, and corticosteroids, and enhanced recovery after surgery protocols. Show less

As the field of implant dentistry continues to evolve, new techniques and technologies arise that can provide great benefits to the partial or completely edentulous patient. The purpose of this article is to review the history, definition, and rationale of immediate loading of dental implants with the goal of providing evidence-based recommendations for implementation into clinical practice. Relevant literature is summarized and includes discussion regarding prerequisites for immediate loading/restoration of an endosseous implant. Surgical techniques and methodologies to prevent implant failure in immediate-load cases are discussed as well. The greatest success has been demonstrated with 4 or more mandibular implants. Although there is support in the literature demonstrating successful outcomes in immediate functional loading of single implants, the opinion of the author is to opt for a nonfunctional load that does not have any occlusal contacts when considering immediate loading of a single dental implant. Show less

This article provides an update for the practicing dentist and/or oral and maxillofacial surgeon on the recognition, identification, and treatment of burning mouth syndrome (BMS). We discuss the most common clinical findings and most common causes of BMS. This article provides a classification flowchart that assists the practitioner in diagnosing and classifying BMS. The article then discusses the pathophysiology and treatment of BMS updated in the literature from the latest studies and reviews. Treatment can vary from topical or systemic medication to behavioral therapy. Show less

Plants have evolved photosynthetic regulatory mechanisms to maintain homeostasis in response to light changes during diurnal transitions and those caused by passing clouds or by wind. One such adaptation directs photosynthetic electron flow to a cyclic pathway to alleviate excess energy surges. Here, we assign a function to regulatory cysteines of PGR5-like protein 1A (PGRL1A), a constituent of the PROTON GRADIENT REGULATION5 (PGR5)-dependent cyclic electron flow (CEF) pathway. During step increases from darkness to low light intensity in Arabidopsis (Arabidopsis thaliana), the intermolecular disulfide of the PGRL1A 59-kDa complex was reduced transiently within seconds to the 28-kDa form. In contrast, step increases from darkness to high light stimulated a stable, partially reduced redox state in PGRL1A. Mutations of 2 cysteines in PGRL1A, Cys82 and Cys183, resulted in a constitutively pseudo-reduced state. The mutant displayed higher proton motive force (PMF) and nonphotochemical quenching (NPQ) than the wild type (WT) and showed altered donor and acceptor dynamic flow around PSI. These changes were found to correspond with the redox state of PGRL1A. Continuous light regimes did not affect mutant growth compared to the WT. However, under fluctuating regimes of high light, the mutant showed better growth than the WT. In contrast, in fluctuating regimes of low light, the mutant displayed a growth penalty that can be attributed to constant stimulation of CEF under low light. Treatment with photosynthetic inhibitors indicated that PGRL1A redox state control depends on the penultimate Fd redox state. Our results showed that redox state changes in PGRL1A are crucial to optimize photosynthesis. Show less

To evaluate whether epidural analgesia is an independent risk factor for OASIS. A population-based cohort study including all women who delivered by spontaneous vaginal delivery or by instrumental delivery beyond 24 weeks gestation was conducted. Deliveries occurred between 1988 and 2016 at a large university tertiary medical center. Women with multiple gestations and those lacking prenatal care were excluded from the analysis. During the study period, 252,542 women delivered at the Soroka University Medical Center and met the inclusion criteria. Of these, 583 (0.23%) were diagnosed with OASIS. Women with OASIS were more likely to be younger, nulliparous, with suspected fetal macrosomia, had higher rates of labor induction and vacuum extraction delivery, higher rates of conceiving after infertility treatments, more advanced gestational age at delivery, higher mean birth weight, higher rates of post-partum hemorrhage and need for blood transfusions. Use of epidural analgesia during pregnancy was significantly high among the OASIS group. Rates of episiotomy were not significantly different between the groups. Using a multimodal logistic regression model, after controlling for vacuum delivery, large for gestational age, nulliparity, gestational age, ethnicity, maternal age, induction of labor, fertility treatments, non-reassuring fetal heart rate and non-progressive second stage of labor, epidural analgesia was found to be significantly associated with OASIS. Epidural analgesia was found to be an independent risk factor for OASIS in our population. Show less

A bacterial phosphotriesterase was employed as an experimental paradigm to examine the effects of multiple factors, such as the molecular constructs, the ligands used during protein expression and purification, the crystallization conditions and the space group, on the visualization of molecular complexes of ligands with a target enzyme. In this case, the ligands used were organophosphates that are fragments of the nerve agents and insecticides on which the enzyme acts as a bioscavenger. 12 crystal structures of various phosphotriesterase constructs obtained by directed evolution were analyzed, with resolutions of up to 1.38 Å. Both apo forms and holo forms, complexed with the organophosphate ligands, were studied. Crystals obtained from three different crystallization conditions, crystallized in four space groups, with and without N-terminal tags, were utilized to investigate the impact of these factors on visualizing the organophosphate complexes of the enzyme. The study revealed that the tags used for protein expression can lodge in the active site and hinder ligand binding. Furthermore, the space group in which the protein crystallizes can significantly impact the visualization of bound ligands. It was also observed that the crystallization precipitants can compete with, and even preclude, ligand binding, leading to false positives or to the incorrect identification of lead drug candidates. One of the co-crystallization conditions enabled the definition of the spaces that accommodate the substituents attached to the P atom of several products of organophosphate substrates after detachment of the leaving group. The crystal structures of the complexes of phosphotriesterase with the organophosphate products reveal similar short interaction distances of the two partially charged O atoms of the P-O bonds with the exposed β-Zn Show less

Peripartum cardiomyopathy (PPCM) is a rare but potentially devastating complication of pregnancy. Although the pathophysiology of PPCM is not fully understood, there are known risk factors for developing PPCM, which are maternal and gestation related. In the first wave of the coronavirus disease 2019 (COVID-19) pandemic, we witnessed an elevated incidence of PPCM among COVID-19 survivors. To present a single-center case series of three patients diagnosed with peripartum cardiomyopathy after recovered from COVID-19 during the index pregnancy. In this single center case study, all patients diagnosed with PPCM at our institute during the examined time frame were included. Electronic medical records were studied. Three patients previously diagnosed with asymptomatic or mildly symptomatic COVID-19 disease during pregnancy presented with PPCM before or shortly after delivery. Patients underwent testing to rule out residual COVID-19 myocarditis, were treated pharmacologically and with wearable defibrillators as needed, and were examined in follow-up 1-9 months after delivery. Residual endothelial damage due to COVID-19 disease, even if originally mild in presentation, could predispose pregnant patients to PPCM and should be considered as a risk factor when assessing patients with new onset symptoms of heart failure. Further research is needed to confirm this hypothesis and fully determine the underlying pathophysiology. These preliminary findings warrant a high index of suspicion for PPCM in COVID-19 recoverers. Show less

Biological aging can be described as accumulative, prolonged metabolic stress and is the major risk factor for cognitive decline and Alzheimer's disease (AD). Recently, we identified and described a quinone reductase 2 (QR2) pathway in the brain, in which QR2 acts as a removable memory constraint and metabolic buffer within neurons. QR2 becomes overexpressed with age, and it is possibly a novel contributing factor to age-related metabolic stress and cognitive deficit. We found that, in human cells, genetic removal of QR2 produced a shift in the proteome opposing that found in AD brains while simultaneously reducing oxidative stress. We therefore created highly specific QR2 inhibitors (QR2is) to enable evaluation of chronic QR2 inhibition as a means to reduce biological age-related metabolic stress and cognitive decline. QR2is replicated results obtained by genetic removal of QR2, while local QR2i microinjection improved hippocampal and cortical-dependent learning in rats and mice. Continuous consumption of QR2is in drinking water improved cognition and reduced pathology in the brains of AD-model mice (5xFAD), with a noticeable between-sex effect on treatment duration. These results demonstrate the importance of QR2 activity and pathway function in the healthy and neurodegenerative brain and what we believe to be the great therapeutic potential of QR2is as first-in-class drugs. Show less

CysD domains are disulfide-rich modules embedded within long O-glycosylated regions of mucin glycoproteins. CysD domains are thought to mediate intermolecular adhesion during the intracellular bioassembly of mucin polymers and perhaps also after secretion in extracellular mucus hydrogels. The human genome encodes 18 CysD domains distributed across three different mucins. To date, experimental structural information is available only for the first CysD domain (CysD1) of the intestinal mucin MUC2, which is one of the most divergent of the CysDs. To provide experimental data on a CysD that is representative of a larger branch of the fold family, we determined the crystal structure of the seventh CysD domain (CysD7) from MUC5AC, a mucin found in the respiratory tract and stomach. The MUC5AC CysD7 structure revealed a single calcium-binding site, contrasting with the two sites in MUC2 CysD1. The MUC5AC CysD7 structure also contained an additional α-helix absent from MUC2 CysD1, with potential functional implications for intermolecular interactions. Lastly, the experimental structure emphasized the flexibility of the loop analogous to the main adhesion loop of MUC2 CysD1, suggesting that both sequence divergence and physical plasticity in this region may contribute to the adaptation of mucin CysD domains. Show less

Phytoplankton produce the volatile dimethyl sulfide (DMS), an important infochemical mediating microbial interactions, which is also emitted to the atmosphere and affecting the global climate. Albeit the enzymatic source for DMS in eukaryotes was elucidated, namely a DMSP lyase (DL) called Alma1, we still lack basic knowledge regarding its taxonomic distribution. We defined unique sequence motifs which enable the identification of DL homologs (DLHs) in model systems and environmental populations. We used these motifs to predict DLHs in diverse algae by analyzing hundreds of genomic and transcriptomic sequences from model systems under stress conditions and from environmental samples. Our findings show that the DL enzyme is more taxonomically widespread than previously thought, as it is encoded by known algal taxa as haptophytes and dinoflagellates, but also by chlorophytes, pelagophytes and diatoms, which were conventionally considered to lack the DL enzyme. By exploring the Tara Oceans database, we showed that DLHs are widespread across the oceans and are predominantly expressed by dinoflagellates. Certain dinoflagellate DLHs were differentially expressed between the euphotic and mesopelagic zones, suggesting a functional specialization and an involvement in the metabolic plasticity of mixotrophic dinoflagellates. In specific regions as the Southern Ocean, DLH expression by haptophytes and diatoms was correlated with environmental drivers such as nutrient availability. The expanded repertoire of putative DL enzymes from diverse microbial origins and geographic niches suggests new potential players in the marine sulfur cycle and provides a foundation to study the cellular function of the DL enzyme in marine microbes. Show less

Albumin is the most abundant protein in the blood serum of mammals and has essential carrier and physiological roles. Albumins are also used in a wide variety of molecular and cellular experiments and in the cultivated meat industry. Despite their importance, however, albumins are challenging for heterologous expression in microbial hosts, likely due to 17 conserved intramolecular disulfide bonds. Therefore, albumins used in research and biotechnological applications either derive from animal serum, despite severe ethical and reproducibility concerns, or from recombinant expression in yeast or rice. We use the PROSS algorithm to stabilize human and bovine serum albumins, finding that all are highly expressed in E. coli. Design accuracy is verified by crystallographic analysis of a human albumin variant with 16 mutations. This albumin variant exhibits ligand binding properties similar to those of the wild type. Remarkably, a design with 73 mutations relative to human albumin exhibits over 40 °C improved stability and is stable beyond the boiling point of water. Our results suggest that proteins with many disulfide bridges have the potential to exhibit extreme stability when subjected to design. The designed albumins may be used to make economical, reproducible, and animal-free reagents for molecular and cell biology. They also open the way to high-throughput screening to study and enhance albumin carrier properties. Show less

Cellular lineage tracking provides a means to observe population makeup at the clonal level, allowing exploration of heterogeneity, evolutionary and developmental processes and individual clones' relative fitness. It has thus contributed significantly to understanding microbial evolution, organ differentiation and cancer heterogeneity, among others. Its use, however, is limited because existing methods are highly specific, expensive, labour-intensive, and, critically, do not allow the repetition of experiments. To address these issues, we developed gUMI-BEAR (genomic Unique Molecular Identifier Barcoded Enriched Associated Regions), a modular, cost-effective method for tracking populations at high resolution. We first demonstrate the system's application and resolution by applying it to track tens of thousands of Saccharomyces cerevisiae lineages growing together under varying environmental conditions applied across multiple generations, revealing fitness differences and lineage-specific adaptations. Then, we demonstrate how gUMI-BEAR can be used to perform parallel screening of a huge number of randomly generated variants of the Hsp82 gene. We further show how our method allows isolation of variants, even if their frequency in the population is low, thus enabling unsupervised identification of modifications that lead to a behaviour of interest. Show less

The increased frequency of mass shootings, terror attacks, and natural disasters in recent years have presented challenges to provision of quality medical care in both short and long-term stressful situations. While emergency departments and trauma surgeons are usually the face of the response to mass casualty incidents (MCI), other departments such as radiology are often active participants in caring for these patients but may not be as well prepared. In this article, we review nine papers describing the experiences of various radiology departments with specific MCIs and the lessons they learned from those experiences. By analysis of common themes raised in these papers, we hope to enable departments to incorporate these lessons into their disaster plans to enhance their preparedness for such events. Show less

The composition, sequence, length and type of glycosidic linkage of polysaccharides profoundly affect their biological and physical properties. However, investigation of the structure-function relationship of polysaccharides is hampered by difficulties in accessing well-defined polysaccharides in sufficient quantities. Here we report a chemical approach to precision polysaccharides with native glycosidic linkages via living cationic ring-opening polymerization of 1,6-anhydrosugars. We synthesized well-defined polysaccharides with tunable molecular weight, low dispersity and excellent regio- and stereo-selectivity using a boron trifluoride etherate catalyst and glycosyl fluoride initiators. Computational studies revealed that the reaction propagated through the monomer α-addition to the oxocarbenium and was controlled by the reversible deactivation of the propagating oxocarbenium to form the glycosyl fluoride dormant species. Our method afforded a facile and scalable pathway to multiple biologically relevant precision polysaccharides, including D-glucan, D-mannan and an unusual L-glucan. We demonstrated that catalytic depolymerization of precision polysaccharides efficiently regenerated monomers, suggesting their potential utility as a class of chemically recyclable materials with tailored thermal and mechanical properties. Show less

Enzymes play a vital role in life processes; they control chemical reactions and allow functional cycles to be synchronized. Many enzymes harness large-scale motions of their domains to achieve tremendous catalytic prowess and high selectivity for specific substrates. One outstanding example is provided by the three-domain enzyme adenylate kinase (AK), which catalyzes phosphotransfer between ATP to AMP. Here we study the phenomenon of substrate inhibition by AMP and its correlation with domain motions. Using single-molecule FRET spectroscopy, we show that AMP does not block access to the ATP binding site, neither by competitive binding to the ATP cognate site nor by directly closing the LID domain. Instead, inhibitory concentrations of AMP lead to a faster and more cooperative domain closure by ATP, leading in turn to an increased population of the closed state. The effect of AMP binding can be modulated through mutations throughout the structure of the enzyme, as shown by the screening of an extensive AK mutant library. The mutation of multiple conserved residues reduces substrate inhibition, suggesting that substrate inhibition is an evolutionary well conserved feature in AK. Combining these insights, we developed a model that explains the complex activity of AK, particularly substrate inhibition, based on the experimentally observed opening and closing rates. Notably, the model indicates that the catalytic power is affected by the microsecond balance between the open and closed states of the enzyme. Our findings highlight the crucial role of protein motions in enzymatic activity. Show less

Acid-β-glucosidase (GCase, EC3.2.1.45), the lysosomal enzyme which hydrolyzes the simple glycosphingolipid, glucosylceramide (GlcCer), is encoded by the GBA1 gene. Biallelic mutations in GBA1 cause the human inherited metabolic disorder, Gaucher disease (GD), in which GlcCer accumulates, while heterozygous GBA1 mutations are the highest genetic risk factor for Parkinson's disease (PD). Recombinant GCase (e.g., Cerezyme Show less

Oxalic acid is a small metabolite found in many plants. It serves as protection from herbivores, a chelator of metal ions, a regulator of calcium levels, and additional tasks. However, it is also a strong di-carboxylic acid that can compromise plant viability by reducing cellular pH. Several metabolic pathways have evolved to control oxalate levels in plants by enzymatic degradation. Among them is the pathway that utilizes oxalyl-CoA synthetase (OCS, EC 6.2.1.8) and ATP to convert oxalate to oxalyl-CoA. Oxalyl-CoA can then be degraded to CO Show less

Grass pea (Lathyrus sativus L.) is a grain legume commonly grown in Asia and Africa for food and forage. It is a highly nutritious and robust crop, capable of surviving both droughts and floods. However, it produces a neurotoxic compound, β-N-oxalyl-L-α,β-diaminopropionic acid (β-ODAP), which can cause a severe neurological disorder when consumed as a primary diet component. While the catalytic activity associated with β-ODAP formation was demonstrated more than 50 years ago, the enzyme responsible for this activity has not been identified. Here, we report on the identity, activity, 3D structure, and phylogenesis of this enzyme-β-ODAP synthase (BOS). We show that BOS belongs to the benzylalcohol O-acetyltransferase, anthocyanin O-hydroxycinnamoyltransferase, anthranilate N-hydroxycinnamoyl/benzoyltransferase, deacetylvindoline 4-O-acetyltransferase superfamily of acyltransferases and is structurally similar to hydroxycinnamoyl transferase. Using molecular docking, we propose a mechanism for its catalytic activity, and using heterologous expression in tobacco leaves (Nicotiana benthamiana), we demonstrate that expression of BOS in the presence of its substrates is sufficient for β-ODAP production in vivo. The identification of BOS may pave the way toward engineering β-ODAP-free grass pea cultivars, which are safe for human and animal consumption. Show less