Compaction, Relaxation, and Linearization of Megadalton-Sized DNA Plasmids: DNA Structures Probed by CD-MS.

High purity plasmid DNA is a raw material for recombinant protein production as well as an active ingredient in DNA vaccines. There are four primary plasmid structures that can be observed in a typical plasmid formulation: supercoiled, relaxed (circular), linearized, and condensed. Determining what structures are present in a sample is important, as the structure can affect activity; the supercoiled structure has the highest activity, and >90% supercoiled is desired for industry standards. Recently, charge detection mass spectrometry (CD-MS) was used to distinguish two of the structures, supercoiled and condensed, by measuring the charge deposited on the ions by positive mode electrospray. Here, CD-MS is used to probe the structures of DNA plasmids during compaction with polycations, and through enzymatic treatment to relax and linearize plasmids. We find that all four structural types for plasmid DNA have unique charging profiles that can be distinguished using CD-MS. The extent of mechanical shearing of the DNA plasmids during electrospray is strongly influenced by the structural type.

Plasma sphingolipids mediate the association between gut microbiome composition and type 2 diabetes risk in the HELIUS cohort: a case-cohort study.

INTRODUCTION: The association between the gut microbiome and incident type 2 diabetes (T2D) is potentially partly mediated through sphingolipids, however these possible mediating mechanisms have not been investigated. We examined whether sphingolipids mediate the association between gut microbiome and T2D, using data from the Healthy Life in an Urban Setting study. RESEARCH DESIGN AND METHODS: Participants were of Dutch or South-Asian Surinamese ethnicity, aged 18-70 years, and without T2D at baseline. A case-cohort design (subcohort n=176, cases incident T2D n=36) was used. The exposure was measured by 16S rRNA sequencing (gut microbiome) and mediator by targeted metabolomics (sphingolipids). Dimensionality reduction was achieved by principle component analysis and Shannon diversity. Cox regression and procrustes analyses were used to assess the association between gut microbiome and T2D and sphingolipids and T2D, and between gut microbiome and sphingolipids, respectively. Mediation was tested familywise using mediation analysis with permutation testing and Bonferroni correction. RESULTS: Our study confirmed associations between gut microbiome and T2D and sphingolipids and T2D. Additionally, we showed that the gut microbiome was associated with sphingolipids. The association between gut microbiome and T2D was partly mediated by a sphingolipid principal component, which represents a dominance of ceramide species over more complex sphingolipids (HR 1.17; 95% CI 1.08 to 1.28; proportional explained 48%), and by Shannon diversity (HR 0.97; 95% CI 0.95 to 0.99; proportional explained 24.8%). CONCLUSIONS: These data suggest that sphingolipids mediate the association between microbiome and T2D risk. Future research is needed to confirm observed findings and elucidate causality on a molecular level.

Pseudo-Worsening of Kidney Function Due to Inhibition of Renal Creatinine Secretion: Quality of Information Provided in Prescribing Information/SmPC.

Determination of serum creatinine concentrations and subsequent calculation of estimated glomerular filtration rates (eGFR) is a cornerstone of clinical medicine. Crucial clinical decisions such as drug treatment discontinuations are based on eGFR calculated from serum creatinine measurements. However, creatinine is not only filtered in the kidneys, but also actively secreted into urine. Creatinine transporters such as OCT2, OCT3, MATE1, MATE2-K, and OAT2 expressed in proximal tubular cells are responsible for active renal secretion of creatinine. Multiple drugs (e.g., oral antitumor drugs) inhibit these transporters thereby causing a pseudo-worsening of kidney function with an increase in serum creatinine concentrations and a decrease in eGFR while other methods for eGFR determination (e.g., by cystatin C) reveal normal kidney function. Since US Prescribing Information (PI) and European Summaries of Product Characteristics (SmPCs) are the most relevant source of information for physicians, we investigated the quality of information in US PI/German SmPCs of drugs with clear evidence for pseudo-worsening of kidney function. 514 drugs putatively interacting with creatinine transporters were identified. For 149 of those drugs, an increase in serum creatinine concentrations has been described. Available data confirmed the existence of pseudo-worsening of kidney function for 30 of those drugs, for the remaining 119 drugs existing data are insufficient. Only 23.5% (12/51) of the 30 drugs' PI/SmPCs contained unambiguous statements on this proven pseudo-worsening of kidney function and gave clear recommendations for clinical management. Taken together, inadequate information provided in PI or SmPCs on the pseudo-worsening of kidney function poses patients at unnecessary risks.

Cell size, density, and nutrient dependency of unicellular algal gravitational sinking velocities.

Eukaryotic phytoplankton, also known as algae, form the basis of marine food webs and drive marine carbon sequestration. Algae must regulate their motility and gravitational sinking to balance access to light at the surface and nutrients in deeper layers. However, the regulation of gravitational sinking remains largely unknown, especially in motile species. Here, we quantify gravitational sinking velocities according to Stokes' law in diverse clades of unicellular marine microalgae to reveal the cell size, density, and nutrient dependency of sinking velocities. We identify a motile algal species, Tetraselmis sp., that sinks faster when starved due to a photosynthesis-driven accumulation of carbohydrates and a loss of intracellular water, both of which increase cell density. Moreover, the regulation of cell sinking velocities is connected to proliferation and can respond to multiple nutrients. Overall, our work elucidates how cell size and density respond to environmental conditions to drive the vertical migration of motile algae.

Food webs in food webs: the micro-macro interplay of multilayered networks.

Food webs are typically defined as being macro-organism-based (e.g., plants, mammals, birds) or microbial (e.g., bacteria, fungi, viruses). However, these characterizations have limits. We propose a multilayered food web conceptual model where microbial food webs are nested within food webs composed of macro-organisms. Nesting occurs through host-microbe interactions, which influence the health and behavior of host macro-organisms, such that host microbiomes likely alter population dynamics of interacting macro-organisms and vice versa. Here, we explore the theoretical underpinnings of multilayered food webs and the implications of this new conceptual model on food web ecology. Our framework opens avenues for new empirical investigations into complex ecological networks and provides a new lens through which to view a network's response to ecosystem changes.

Behavioural activation for people in custody with depression: A protocol for a feasibility randomised controlled study.

People in custody are at high risk of developing depression. Accessing psychological treatments in a prison setting is a particular challenge, in part, due to difficulties accessing specialist mental health workers. Behavioural Activation (BA) may be helpful in improving health outcomes for people in custody experiencing depressive symptoms. The aim of this study is to establish the feasibility and acceptability of custodial health nurses delivering BA to improve depressive symptoms of people in custody. We will conduct a pilot randomised controlled trial with process observation examining the feasibility and acceptability of BA in treating people in custody with depressive symptoms. 60 people in custody presenting with depressive symptoms will be randomised to receive BA plus treatment as usual (TAU) or TAU provided by custodial health nurses. Eight custodial health nurses will be recruited, trained, and deliver BA. BA will be delivered twice a week for six weeks, with sessions lasting up to 30 minutes. Changes in depression and quality of life (QoL) will be assessed at baseline, 6 weeks, and 3 months post-intervention. Participants will be interviewed to understand feasibility and acceptability of BA in prison settings. The findings will inform the design of a randomised controlled trial to test the efficacy of BA for people in custody with depression. Findings will help determine whether BA for depression is suited to prison health care system and services. Improving depressive symptoms in people in custody has benefits beyond prison settings. The Central Adelaide Local Health Network Human Research Ethics Committee and University of South Australia Human Research Ethics Committee have approved the study. The trial results will be disseminated through peer-reviewed journals and scientific conferences and reported to local stakeholders and policy makers. If feasibility and acceptability is demonstrated, we will seek to progress to an effectiveness study. A potential strength of the trial model proposed, is in its scalability, with potential to increase the trial sites and locations. This trial has been prospectively registered with the Australian New Zealand Clinical Trials Registry (reference number: ACTRN12623000346673p). Trial registration ACTRN12623000346673p.

The InBIO Barcoding Initiative Database: DNA barcodes of Portuguese moths.

Background: The InBIO Barcoding Initiative (IBI) Dataset - DS-IBILP08 contains records of 2350 specimens of moths (Lepidoptera species that do not belong to the superfamily Papilionoidea). All specimens have been morphologically identified to species or subspecies level and represent 1158 species in total. The species of this dataset correspond to about 42% of mainland Portuguese Lepidoptera species. All specimens were collected in mainland Portugal between 2001 and 2022. All DNA extracts and over 96% of the specimens are deposited in the IBI collection at CIBIO, Research Center in Biodiversity and Genetic Resources. New information: The authors enabled "The InBIO Barcoding Initiative Database: DNA barcodes of Portuguese moths" in order to release the majority of data of DNA barcodes of Portuguese moths within the InBIO Barcoding Initiative. This dataset increases the knowledge on the DNA barcodes of 1158 species from Portugal belonging to 51 families. There is an increase in DNA barcodes of 205% in Portuguese specimens publicly available. The dataset includes 61 new Barcode Index Numbers. All specimens have their DNA barcodes publicly accessible through BOLD online database and the distribution data can be accessed through the Global Biodiversity Information Facility (GBIF).

Bulk-suppressed and surface-sensitive Raman scattering by transferable plasmonic membranes with irregular slot-shaped nanopores.

Raman spectroscopy enables the non-destructive characterization of chemical composition, crystallinity, defects, or strain in countless materials. However, the Raman response of surfaces or thin films is often weak and obscured by dominant bulk signals. Here we overcome this limitation by placing a transferable porous gold membrane, (PAuM) on the surface of interest. Slot-shaped nanopores in the membrane act as plasmonic antennas and enhance the Raman response of the surface or thin film underneath. Simultaneously, the PAuM suppresses the penetration of the excitation laser into the bulk, efficiently blocking its Raman signal. Using graphene as a model surface, we show that this method increases the surface-to-bulk Raman signal ratio by three orders of magnitude. We find that 90% of the Raman enhancement occurs within the top 2.5 nm of the material, demonstrating truly surface-sensitive Raman scattering. To validate our approach, we quantify the strain in a 12.5 nm thin Silicon film and analyze the surface of a LaNiO3 thin film. We observe a Raman mode splitting for the LaNiO3 surface-layer, which is spectroscopic evidence that the surface structure differs from the bulk. These results validate that PAuM gives direct access to Raman signatures of thin films and surfaces.

The "Twin Peaks" method of automated Spike-Wave detection: A two-step, two-criteria Matlab application.

BACKGROUND: There are many automated spike-wave discharge detectors, but the known weaknesses of otherwise good methods and the varying working conditions of different research groups (mainly the access to hardware and software) invite further exploration into alternative approaches. NEW METHOD: The algorithm combines two criteria, one in the time-domain and one in the frequency-domain, exploiting morphological asymmetry and the presence of harmonics, respectively. The time-domain criterion is additionally adjusted by normal modelling between the first and second iterations. RESULTS: We report specificity, sensitivity and accuracy values for 20 recordings from 17 mature, male WAG/Rij rats. In addition, performance was preliminary tested with different hormones, pharmacological injections and species (mice) in a smaller sample. Accuracy and specificity were consistently above 91 %. The number of automatically detected spike-wave discharges was strongly correlated with the numbers derived from visual inspection. Sensitivity varied more strongly than specificity, but high values were observed in both rats and mice. COMPARISON WITH EXISTING METHODS: The algorithm avoids low-voltage movement artifacts, displays a lower false positive rate than many predecessors and appears to work across species, i.e. while designed initially with data from the WAG/Rij rat, the algorithm can pick up seizure activity in the mouse of considerably lower inter-spike frequency. Weaknesses of the proposed method include a lower sensitivity than several predecessors. CONCLUSION: The algorithm excels in being a selective and flexible (based on e.g. its performance across rats and mice) spike-wave discharge detector. Future work could attempt to increase the sensitivity of this approach.

Dietary fibre directs microbial tryptophan metabolism via metabolic interactions in the gut microbiota.

Tryptophan is catabolized by gut microorganisms resulting in a wide range of metabolites implicated in both beneficial and adverse host effects. How gut microbial tryptophan metabolism is directed towards indole, associated with chronic kidney disease, or towards protective indolelactic acid (ILA) and indolepropionic acid (IPA) is unclear. Here we used in vitro culturing and animal experiments to assess gut microbial competition for tryptophan and the resulting metabolites in a controlled three-species defined community and in complex undefined human faecal communities. The generation of specific tryptophan-derived metabolites was not predominantly determined by the abundance of tryptophan-metabolizing bacteria, but rather by substrate-dependent regulation of specific metabolic pathways. Indole-producing Escherichia coli and ILA- and IPA-producing Clostridium sporogenes competed for tryptophan within the three-species community in vitro and in vivo. Importantly, fibre-degrading Bacteroides thetaiotaomicron affected this competition by cross-feeding monosaccharides to E. coli. This inhibited indole production through catabolite repression, thus making more tryptophan available to C. sporogenes, resulting in increased ILA and IPA production. The fibre-dependent reduction in indole was confirmed using human faecal cultures and faecal-microbiota-transplanted gnotobiotic mice. Our findings explain why consumption of fermentable fibres suppresses indole production but promotes the generation of other tryptophan metabolites associated with health benefits.

The Conservative and Operative Treatment of Carpal Fractures.

BACKGROUND: Carpal fractures (incidence: 30-60 per 100 000 persons per year) are one of the more commonly overlooked fracture types. They can have serious consequences, as the use of the hand is indispensable in everyday life. In the following article, we present the elements of the diagnosis and treatment of fractures of the carpal bones. METHODS: This review is based on meta-analyses and randomized controlled trials (RCTs) published from 2013 to 2023 that were retrieved by a structured literature search, supplemented by guideline recommendations and expert consensus statements. In addition, data on the administrative prevalence of carpal fractures were obtained from the German Association of Statutory Health Insurance Physicians (Kassenärztliche Vereinigung, KV) and from the German Statutory Accident Insurance (Deutsche Gesetzliche Unfallversicherung, DGUV). RESULTS: The administrative prevalence of carpal fractures in 2022 was 44 496 outpatient cases (KV, DGUV) in one year. After clinical history-taking, physical examination and x-ray have been performed, thin-slice computed tomography is recommended as part of the diagnostic evaluation. Treatment recommendations are based on evidence of levels II to IV. Multiple RCTs have been carried out on the treatment of scaphoid fractures, and a clinical guideline exists. Proximal, dislocated and unstable scaphoid fractures should be treated surgically. Non-displaced or minimally displaced fractures of the middle third of the scaphoid bone require a shorter period of immobilization with surgical treatment (2-4 weeks) than with conservative treatment (6-8 weeks). The use of plaster casts that do not hinder elbow and thumb mobility yields healing rates similar to those obtained with the immobilization of both of these joints. Failure to treat an unrecognized scaphoid fracture can lead to pseudarthrosis, avascular bony necrosis, and misalignment. Other, rarer types of carpal fractures must be managed on an individual basis, as the available evidence is limited to expert consensus. CONCLUSION: Early recognition and appropriate treatment of carpal fractures lead to healing in more than 90% of cases. Although the available evidence on their proper treatment is growing, many questions are subject to expert consensus, and decisions about treatment must be made individually.

Identifying Key Drivers of Efficient B Cell Responses: On the Role of T Help, Antigen-Organization, and Toll-like Receptor Stimulation for Generating a Neutralizing Anti-Dengue Virus Response.

T help (Th), stimulation of toll-like receptors (pathogen-associated molecular patterns, PAMPs), and antigen organization and repetitiveness (pathogen-associated structural patterns, PASPs) were shown numerous times to be important in driving B-cell and antibody responses. In this study, we dissected the individual contributions of these parameters using newly developed "Immune-tag" technology. As model antigens, we used eGFP and the third domain of the dengue virus 1 envelope protein (DV1 EDIII), the major target of virus-neutralizing antibodies. The respective proteins were expressed alone or genetically fused to the N-terminal fragment of the cucumber mosaic virus (CMV) capsid protein-nCMV, rendering the antigens oligomeric. In a step-by-step manner, RNA was attached as a PAMP, and/or a universal Th-cell epitope was genetically added for additional Th. Finally, a PASP was added to the constructs by displaying the antigens highly organized and repetitively on the surface of CMV-derived virus-like particles (CuMV VLPs). Sera from immunized mice demonstrated that each component contributed stepwise to the immunogenicity of both proteins. All components combined in the CuMV VLP platform induced by far the highest antibody responses. In addition, the DV1 EDIII induced high levels of DENV-1-neutralizing antibodies only if displayed on VLPs. Thus, combining multiple cues typically associated with viruses results in optimal antibody responses.

A Hybrid Type III Effectiveness-Implementation Trial to Optimize Medication Safety With Oral Antitumor Therapy in Real-World: The AMBORA Competence and Consultation Center.

PURPOSE: Implementation science endeavors to facilitate the translation of evidence-based research into clinical routine. The clinical pharmacological/pharmaceutical care program evaluated in the randomized AMBORA trial on medication safety with oral antitumor therapeutics (OAT) optimizes care delivery and provides significant benefits for patients, treatment teams, and health care systems. Thus, we aimed to investigate the implementation of this care program within the AMBORA Competence and Consultation Center (AMBORA Center). METHODS: The AMBORA Center within a University Comprehensive Cancer Center offered several services (eg, patient consultations) and was evaluated according to the RE-AIM framework. This multicenter hybrid type III trial focused on implementation outcomes (eg, patient recruitment, referring units, evaluation of services) while concurrently investigating effectiveness (eg, side effects, medication errors). Quantitative and qualitative assessments were combined. RESULTS: The AMBORA Center conducted over 800 consultations with 420 patients in seven institutions. The primary end point of counseling 70% of patients treated with OAT was not reached. Patients were referred by 15 treatment units compared with 11 units in the AMBORA trial. On the basis of heterogeneous referral rates and characteristics across the institutions, barriers and facilitators of the implementation process were derived. Several survey results (eg, stakeholder interviews, online/paper-based questionnaires) reflected a high appreciation of services by patients and health care professionals. The severity of 60.1% (178 of 296) of detected side effects improved, and 86.3% (297 of 344) of medication errors were resolved. CONCLUSION: Despite not reaching the primary implementation outcome, the AMBORA Center included more treatment units and demonstrated patient benefit of the AMBORA care program by meeting all effectiveness outcomes. We outlined quantitative and qualitative implementation characteristics to enhance outreach and foster further dissemination of centers to optimize medication safety with OAT.

The median nerve´s system of connective tissue distal to the pronator teres to the carpal tunnel.

BACKGROUND: Connective tissue serves a role beyond mere spatial filling. Furthermore, there is increasing evidence that connective tissue plays an important role in the pathogenesis of conditions such as carpal tunnel syndrome (CTS). According to our hypothesis, the median nerve (MN) is surrounded by a system of connective tissue distal to the pronator teres and extending up to, and including, the carpal tunnel. METHODS: To visualize the connective tissue surrounding the median nerve, we dissected the forearms of 15 body donors from pronator teres to the carpal tunnel, created plastination slices stained with Periodic Acid-Schiff (PAS), and injected ink into the seen spaces. We verified our findings with a segmentational analysis of radiological data of 10 healthy individuals. RESULTS: We macroscopically describe the median nerve´s system of connective tissue (MC) distal to the pronator teres and up to and including the carpal tunnel. This system creates, connects, and separates spaces. At least from the pronator teres to the carpal tunnel it also creates subspaces from proximal to distal. For the MC, we established a mean cross-sectional area of 153.1 mm2 (SD=37.15) in the carpal tunnel. The median nerve consistently resides at the center of this MC, which further connects to flexor muscles of the forearm, and to the radius bone. In the carpal tunnel, the MC creates subspaces inside. There, it also acts as the outermost internal layer enveloping flexor tendons, and the MN. DISCUSSION: The term MC does not negate but orders the existence of other "connectives", like subsynovial connective tissue, endo-, epi- or perineuria, epimysia, periostea, or peritendinea, to a hierarchy related to the median nerve. Diseases of the MN are common. Knowing the anatomy of the MC and how it relates to MN function may help clinicians recognize and understand conditions like CTS.

Generation of Rapid and High-Quality Serum by Recombinant Prothrombin Activator Ecarin (RAPClot™).

We recently reported the potential application of recombinant prothrombin activator ecarin (RAPClot™) in blood diagnostics. In a new study, we describe RAPClot™ as an additive to develop a novel blood collection prototype tube that produces the highest quality serum for accurate biochemical analyte determination. The drying process of the RAPClot™ tube generated minimal effect on the enzymatic activity of the prothrombin activator. According to the bioassays of thrombin activity and plasma clotting, γ-radiation (>25 kGy) resulted in a 30-40% loss of the enzymatic activity of the RAPClot™ tubes. However, a visual blood clotting assay revealed that the γ-radiation-sterilized RAPClot™ tubes showed a high capacity for clotting high-dose heparinized blood (8 U/mL) within 5 min. This was confirmed using Thrombelastography (TEG), indicating full clotting efficiency under anticoagulant conditions. The storage of the RAPClot™ tubes at room temperature (RT) for greater than 12 months resulted in the retention of efficient and effective clotting activity for heparinized blood in 342 s. Furthermore, the enzymatic activity of the RAPClot™ tubes sterilized with an electron-beam (EB) was significantly greater than that with γ-radiation. The EB-sterilized RAPClot™ tubes stored at RT for 251 days retained over 70% enzyme activity and clotted the heparinized blood in 340 s after 682 days. Preliminary clinical studies revealed in the two trials that 5 common analytes (K, Glu, lactate dehydrogenase (LD), Fe, and Phos) or 33 analytes determined in the second study in the γ-sterilized RAPClot™ tubes were similar to those in commercial tubes. In conclusion, the findings indicate that the novel RAPClot™ blood collection prototype tube has a significant advantage over current serum or lithium heparin plasma tubes for routine use in measuring biochemical analytes, confirming a promising application of RAPClot™ in clinical medicine.

Ataxia Telangiectasia patient-derived neuronal and brain organoid models reveal mitochondrial dysfunction and oxidative stress.

Ataxia Telangiectasia (AT) is a rare disorder caused by mutations in the ATM gene and results in progressive neurodegeneration for reasons that remain poorly understood. In addition to its central role in nuclear DNA repair, ATM operates outside the nucleus to regulate metabolism, redox homeostasis and mitochondrial function. However, a systematic investigation into how and when loss of ATM affects these parameters in relevant human neuronal models of AT was lacking. We therefore used cortical neurons and brain organoids from AT-patient iPSC and gene corrected isogenic controls to reveal levels of mitochondrial dysfunction, oxidative stress, and senescence that vary with developmental maturity. Transcriptome analyses identified disruptions in regulatory networks related to mitochondrial function and maintenance, including alterations in the PARP/SIRT signalling axis and dysregulation of key mitophagy and mitochondrial fission-fusion processes. We further show that antioxidants reduce ROS and restore neurite branching in AT neuronal cultures, and ameliorate impaired neuronal activity in AT brain organoids. We conclude that progressive mitochondrial dysfunction and aberrant ROS production are important contributors to neurodegeneration in AT and are strongly linked to ATM's role in mitochondrial homeostasis regulation.

Association between claims-based setting of diagnosis and treatment initiation among Medicare patients with hepatitis C.

OBJECTIVE: To develop a claims-based algorithm to determine the setting of a disease diagnosis. DATA SOURCES AND STUDY SETTING: Medicare enrollment and claims data from 2014 to 2019. STUDY DESIGN: We developed a claims-based algorithm using facility indicators, revenue center codes, and place of service codes to identify settings where HCV diagnosis first appeared. When the first appearance was in a laboratory, we attempted to associate HCV diagnoses with subsequent clinical visits. Face validity was assessed by examining association of claims-based diagnostic settings with treatment initiation. DATA COLLECTION/EXTRACTION METHODS: Patients newly diagnosed with HCV and continuously enrolled in traditional Medicare Parts A, B, and D (12 months before and 6 months after index diagnosis) were included. PRINCIPAL FINDINGS: Among 104,454 patients aged 18-64 and 66,726 aged ≥65, 70.1% and 69%, respectively, were diagnosed in outpatient settings, and 20.2% and 22.7%, respectively in laboratory or unknown settings. Logistic regression revealed significantly lower odds of treatment initiation after diagnosis in emergency departments/urgent cares, hospitals, laboratories, or unclassified settings, than in outpatient visits. CONCLUSIONS: The algorithm identified the setting of HCV diagnosis in most cases, and found significant associations with treatment initiation, suggesting an approach that can be adapted for future claims-based studies.

Bortezomib Inhibits Open Configurations of the 20S Proteasome.

Bortezomib, a small dipeptide-like molecule, is a proteasome inhibitor used widely in the treatment of myeloma and lymphoma. This molecule reacts with threonine side chains near the center of the 20S proteasome and disrupts proteostasis by blocking enzymatic sites that are responsible for protein degradation. In this work, we use novel mass-spectrometry-based techniques to examine the influence of bortezomib on the structures and stabilities of the 20S core particle. These studies indicate that bortezomib binding dramatically favors compact 20S structures (in which the axial gate is closed) over larger structures (in which the axial gate is open)─suppressing gate opening by factors of at least ∼400 to 1300 over the temperature range that is studied. Thus, bortezomib may also restrict degradation in the 20S proteasome by preventing substrates from entering the catalytic pore. That bortezomib influences structures at the entrance region of the pore at such a long distance (∼65 to 75 Å) from its binding sites raises a number of interesting biophysical issues.

Bioenergetic mapping of 'healthy microbiomes' via compound processing potential imprinted in gut and soil metagenomes.

Despite mounting evidence of their importance in human health and ecosystem functioning, the definition and measurement of 'healthy microbiomes' remain unclear. More advanced knowledge exists on health associations for compounds used or produced by microbes. Environmental microbiome exposures (especially via soils) also help shape, and may supplement, the functional capacity of human microbiomes. Given the synchronous interaction between microbes, their feedstocks, and micro-environments, with functional genes facilitating chemical transformations, our objective was to examine microbiomes in terms of their capacity to process compounds relevant to human health. Here we integrate functional genomics and biochemistry frameworks to derive new quantitative measures of in silico potential for human gut and environmental soil metagenomes to process a panel of major compound classes (e.g., lipids, carbohydrates) and selected biomolecules (e.g., vitamins, short-chain fatty acids) linked to human health. Metagenome functional potential profile data were translated into a universal compound mapping 'landscape' based on bioenergetic van Krevelen mapping of function-level meta-compounds and corresponding functional relative abundances, reflecting imprinted genetic capacity of microbiomes to metabolize an array of different compounds. We show that measures of 'compound processing potential' associated with human health and disease (examining atherosclerotic cardiovascular disease, colorectal cancer, type 2 diabetes and anxious-depressive behavior case studies), and displayed seemingly predictable shifts along gradients of ecological disturbance in plant-soil ecosystems (three case studies). Ecosystem quality explained 60-92 % of variation in soil metagenome compound processing potential measures in a post-mining restoration case study dataset. With growing knowledge of the varying proficiency of environmental microbiota to process human health associated compounds, we might design environmental interventions or nature prescriptions to modulate our exposures, thereby advancing microbiota-oriented approaches to human health. Compound processing potential offers a simplified, integrative approach for applying metagenomics in ongoing efforts to understand and quantify the role of microbiota in environmental- and human-health.

Point mutation in a virus-like capsid drives symmetry reduction to form tetrahedral cages.

Protein capsids are a widespread form of compartmentalization in nature. Icosahedral symmetry is ubiquitous in capsids derived from spherical viruses, as this geometry maximizes the internal volume that can be enclosed within. Despite the strong preference for icosahedral symmetry, we show that simple point mutations in a virus-like capsid can drive the assembly of unique symmetry-reduced structures. Starting with the encapsulin from Myxococcus xanthus, a 180-mer bacterial capsid that adopts the well-studied viral HK97 fold, we use mass photometry and native charge detection mass spectrometry to identify a triple histidine point mutant that forms smaller dimorphic assemblies. Using cryoelectron microscopy, we determine the structures of a precedented 60-mer icosahedral assembly and an unexpected 36-mer tetrahedron that features significant geometric rearrangements around a new interaction surface between capsid protomers. We subsequently find that the tetrahedral assembly can be generated by triple-point mutation to various amino acids and that even a single histidine point mutation is sufficient to form tetrahedra. These findings represent a unique example of tetrahedral geometry when surveying all characterized encapsulins, HK97-like capsids, or indeed any virus-derived capsids reported in the Protein Data Bank, revealing the surprising plasticity of capsid self-assembly that can be accessed through minimal changes in the protein sequence.