The cornerstone of autotrophy, the CO₂-fixing enzyme, D-ribulose1,5-bisphosphate carboxylase/oxygenase (Rubisco), is hamstrung by slow catalysis and confusion between CO₂ and O₂ as substrates, an ..."abominably perplexing" puzzle, in Darwin's parlance. Here we argue that these characteristics stem from difficulty in binding the featureless CO₂ molecule, which forces specificity for the gaseous substrate to be determined largely or completely in the transition state. We hypothesize that natural selection for greater CO₂/O₂ specificity, in response to reducing atmospheric CO₂:O₂ ratios, has resulted in a transition state for CO₂ addition in which the CO₂ moiety closely resembles a carboxylate group. This maximizes the structural difference between the transition states for carboxylation and the competing oxygenation, allowing better differentiation between them. However, increasing structural similarity between the carboxylation transition state and its carboxyketone product exposes the carboxyketone to the strong binding required to stabilize the transition state and causes the carboxyketone intermediate to bind so tightly that its cleavage to products is slowed. We assert that all Rubiscos may be nearly perfectly adapted to the differing CO₂, O₂, and thermal conditions in their subcellular environments, optimizing this compromise between CO₂/O₂ specificity and the maximum rate of catalytic turnover. Our hypothesis explains the feeble rate enhancement displayed by Rubisco in processing the exogenously supplied carboxyketone intermediate, compared with its nonenzymatic hydrolysis, and the positive correlation between CO₂/O₂ specificity and $^{12}C/^{13}C$ fractionation. It further predicts that, because a more product-like transition state is more ordered (decreased entropy), the effectiveness of this strategy will deteriorate with increasing temperature.
The maintenance of chromosome termini, or telomeres, requires the action of the enzyme telomerase, as conventional DNA polymerases cannot fully replicate the ends of linear molecules. Telomerase is ...expressed and telomere length is maintained in human germ cells and the great majority of primary human tumours. However, telomerase is not detectable in most normal somatic cells; this corresponds to the gradual telomere loss observed with each cell division. It has been proposed that telomere erosion eventually signals entry into senescence or cell crisis and that activation of telomerase is usually required for immortal cell proliferation. In addition to the human telomerase RNA component (hTR; ref. 11), TR1/TLP1 (refs 12, 13), a protein that is homologous to the p80 protein associated with the Tetrahymena enzyme, has been identified in humans. More recently, the human telomerase reverse transcriptase (hTRT; refs 15, 16), which is homologous to the reverse transcriptase (RT)-like proteins associated with the Euplotes aediculatus (Ea_p123), Saccharomyces cerevisiae (Est2p) and Schizosaccharomyces pombe (5pTrt1) telomerases, has been reported to be a telomerase protein subunit. A catalytic function has been demonstrated for Est2p in the RT-like class but not for p80 or its homologues. We now report that in vitro transcription and translation of hTRT when co-synthesized or mixed with hTR reconstitutes telomerase activity that exhibits enzymatic properties like those of the native enzyme. Single amino-acid changes in conserved telomerase-specific and RT motifs reduce or abolish activity, providing direct evidence that hTRT is the catalytic protein component of telomerase. Normal human diploid cells transiently expressing hTRT possessed telomerase activity, demonstrating that hTRT is the limiting component necessary for restoration of telomerase activity in these cells. The ability to reconstitute telomerase permits further analysis of its biochemical and biological roles in cell aging and carcinogenesis.
Catalytic protein subunits of telomerase from the ciliate Euplotes aediculatus and the yeast Saccharomyces cerevisiae contain reverse transcriptase motifs. Here the homologous genes from the fission ...yeast Schizosaccharomyces pombe and human are identifed. Disruption of the S. pombe gene resulted in telomere shortening and senescence, and expression of mRNA from the human gene correlated with telomerase activity in cell lines. Sequence comparisons placed the telomerase proteins in the reverse transcriptase family but revealed hallmarks that distinguish them from retroviral and retrotransposon relatives. Thus, the proposed telomerase catalytic subunits are phylogenetically conserved and represent a deep branch in the evolution of reverse transcriptases
Abstract
We augment the heliospheric network of galactic cosmic ray (GCR) monitors using 2012–2017 penetrating radiation measurements from the New Horizons (NH) Pluto Energetic Particle Spectrometer ...Science Investigation (PEPSSI), obtaining intensities of ≳75 MeV particles. The new, predominantly GCR observations provide critical links between the Sun and Voyager 2 and Voyager 1 (V2 and V1), in the heliosheath and local interstellar medium (LISM), respectively. We provide NH, Advanced Composition Explorer (ACE), V2, and V1 GCR observations, using them to track solar cycle variations and short-term Forbush decreases from the Sun to the LISM, and to examine the interaction that results in the surprising, previously reported V1 LISM anisotropy episodes. To investigate these episodes and the hitherto unexplained lagging of associated in situ shock features at V1, propagating disturbances seen at ACE, NH, and V2 were compared to V1. We conclude that the region where LISM magnetic field lines drape around the heliopause is likely critical for communicating solar disturbance signals upstream of the heliosheath to V1. We propose that the anisotropy-causing physical process that suppresses intensities at ∼90° pitch angles relies on GCRs escaping from a single compression in the draping region, not on GCRs trapped between two compressions. We also show that NH suprathermal and energetic particle data from PEPSSI are consistent with the interpretation that traveling shocks and corotating interaction region (CIR) remnants can be distinguished by the existence or lack of Forbush decreases, respectively, because turbulent magnetic fields at local shocks inhibit GCR transport while older CIR structures reaching the outer heliosphere do not.
Studies evaluating sputum quality and Xpert
MTB/RIF positivity in the context of active case finding are scarce. We aimed to determine whether sputum quality is associated with Xpert positivity and ...whether the association differed according to demographic and clinical characteristics.
A cross-sectional analysis using data from a mass screening programme in Brazilian prisons was conducted from 2017 to 2021. We administered a standardised questionnaire, obtained a chest X-ray and collected a spot sputum sample for Xpert testing. Sputum quality was classified as 'salivary', 'mucoid/mucopurulent' or 'blood-stained'. We used log binomial regressions to estimate the relationship between sputum quality and Xpert positivity, assessing interactions with participant characteristics.
Among 4,368 participants for whom sputum quality was assessed, 957 (21.9%) produced salivary specimens, 3,379 (77.4%) had mucoid/mucopurulent sputum and 32 (0.7%) had blood-stained sputum. Xpert positivity was higher among those with mucoid/mucopurulent sputum than among those with salivary samples (12.0% vs. 3.7%). Mucopurulent sputum independently predicted Xpert positivity among individuals with and without symptoms, current smoking and abnormal chest radiographs on CAD4TB.
In our study, sputum appearance independently predicted Xpert positivity, and could be used together with chest X-ray and symptom screening to inform use of Xpert in individual or pooled testing.
β-Lactam antibiotics disrupt the assembly of peptidoglycan (PG) within the bacterial cell wall by inhibiting the enzymatic activity of penicillin-binding proteins (PBPs). It was recently shown that ...β-lactam treatment initializes a futile cycle of PG synthesis and degradation, highlighting major gaps in our understanding of the lethal effects of PBP inhibition by β-lactam antibiotics. Here, we assess the downstream metabolic consequences of treatment of Escherichia coli with the β-lactam mecillinam and show that lethality from PBP2 inhibition is a specific consequence of toxic metabolic shifts induced by energy demand from multiple catabolic and anabolic processes, including accelerated protein synthesis downstream of PG futile cycling. Resource allocation into these processes is coincident with alterations in ATP synthesis and utilization, as well as a broadly dysregulated cellular redox environment. These results indicate that the disruption of normal anabolic-catabolic homeostasis by PBP inhibition is an essential factor for β-lactam antibiotic lethality.
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•PBP2 inhibition by mecillinam alters bacterial catabolic and anabolic processes•Bacterial protein synthesis is identified as a source of energy demand•Reactive metabolic by-products are important for mecillinam lethality•Target-proximal and downstream metabolic effects contribute to mecillinam lethality
Lobritz et al. assess the target-proximal and downstream metabolic consequences of treatment of E. coli with the β-lactam mecillinam. They show that lethality from PBP2 inhibition is a specific consequence of toxic metabolic shifts induced by energy demand from multiple catabolic and anabolic processes.
Energetic neutral atom imaging instruments have been flown on a variety of space missions to satisfy a variety of science requirements. In this paper we discuss the most recent developments that lead ...to improvements in energy range, angular resolution, and background rejection for the high‐energy range, as represented in the past by the Cassini magnetosphere imaging instrument Ion and Neutral Camera, the Imager for Magnetopause‐to‐Aurora Global Exploration (IMAGE) mission High‐Energy Neutral Atom instrument, and to some degree the IMAGE mission Medium‐Energy Neutral Atom instrument. The new approaches discussed here rely on the use of ultrathin foils without UV filters and on very high speed coincidence logic to reduce accidentals from intense background sources. We present laboratory results demonstrating an electrostatic design that meets the coincidence timing requirements, position, and scattering performance consistent with angular resolution (full width at half maximum) of ~2° for hydrogen above 10 keV and a hydrogen energy threshold ≤1 keV.
Key Points
Energetic neutral atom imaging can be done without filtering ambient ultraviolet light
Angular resolution of 2° and energy range to below 1 keV for hydrogen can be achieved at higher sensitivity than previously
Background rejection sufficient to operate in Jupiter's magnetosphere in the vicinity of Ganymede is possible
This study examines the validity of using ICD-10 codes to identify hospitalized pneumonia cases. Using a case-cohort design, subjects were randomly selected from monthly cohorts of patients aged ⩾65 ...years discharged from April 2000 to March 2002 from two large tertiary Australian hospitals. Cases had ICD-10-AM codes J10–J18 (pneumonia); the cohort sample was randomly selected from all discharges, frequency matched to cases by month. Codes were validated against three comparators: medical record notation of pneumonia, chest radiograph (CXR) report and both. Notation of pneumonia was determined for 5098/5101 eligible patients, and CXR reports reviewed for 3349/3464 (97%) patients with a CXR. Coding performed best against notation of pneumonia: kappa 0·95, sensitivity 97·8% (95% CI 97·1–98·3), specificity 96·9% (95% CI 96·2–97·5), positive predictive value (PPV) 96·2% (95% CI 95·4–97·0) and negative predictive value (NPV) 98·2% (95% CI 97·6–98·6). When medical record notation of pneumonia is used as the standard, ICD-10 codes are a valid method for retrospective ascertainment of hospitalized pneumonia cases and appear superior to use of complexes of symptoms and signs, or radiology reports.
The first orbit of Solar Orbiter provided comprehensive measurements of six corotating interaction regions (CIRs) within 1 au. Five of these CIRs were also observed by ACE at 1 au, allowing for ...comparisons of the suprathermal ion intensities and spectra at different radial distances. Only subtle modulations of the
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He spectral slopes are observed between Solar Orbiter and ACE. Additionally, the radial gradients of 226−320 keV/nuc
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He ion intensities between Solar Orbiter and ACE are similar to that of 1.53 MeV H reported by Van Hollebeke et al. (1978, J. Geophys. Res., 83, A10). These observations provide a new addition to the study of the radial dependence of CIR-associated suprathermal ions in the inner heliosphere.
The objective was to analyze enhancement characteristics of insulinomas and to determine the ability of multiphase CT to localize these tumors.
Prospective interpretations of multiphase helical CT ...scans were reviewed in 30 patients who had insulinomas resected over a 5-year period. CT scans were retrospectively reviewed to determine enhancement characteristics, tumor conspicuity in each phase of enhancement, and potential causes for false-negative findings.
Sixty-three percent (19/30) of tumors were identified on CT prospectively. An additional six tumors were visualized in retrospect, allowing characterization of 25 (83%) of 30 tumors. Most tumors were hyperdense on at least one phase (n = 19), three tumors were hypoattenuating, and three were isodense and pedunculated. Insulinomas were most conspicuous on the early phase in 15 patients and in the portal venous phase in three. All tumors that underwent pancreatic phase imaging were seen (13/13), whereas three of 18 arterial and six of 25 portal venous phase findings were inconclusive for tumor. In the six examinations with false-negative findings in which the tumor could be seen in retrospect, two tumors were isodense and pedunculated, three were in close proximity to vessels, and one had a cystic appearance.
Multiphasic CT has a moderate sensitivity in the detection of insulinomas. Most tumors are more conspicuous on the earlier phases of enhancement. The pancreatic phase may be more useful than the arterial phase. Potential sources of false-negative results include tumors adjacent to vessels, pedunculated morphology, or nonhyperattenuating lesions.
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