Interrogating fragment libraries by X-ray crystallography is a powerful strategy for discovering allosteric ligands for protein targets. Cryocooling of crystals should theoretically increase the fraction of occupied binding sites and decrease radiation damage. However, it might also perturb protein conformations that can be accessed at room temperature. Using data from crystals measured consecutively at room temperature and at cryogenic temperature, we found that transient binding sites could be abolished at the cryogenic temperatures employed by standard approaches. Changing the temperature at which the crystallographic data was collected could provide a deliberate perturbation to the equilibrium of protein conformations and help to visualize hidden sites with great potential to allosterically modulate protein function. Too cool to show up: The common practice of cryocooling crystals was found to mask transient protein conformations. Data collected on a single crystal revealed ligand binding to a cryptic site at room temperature that was hidden at cryogenic temperature. These results contradict thermodynamic expectations and provide a method for discovering allosteric sites with great potential for modulating protein (mal)-function.

A key challenge in structure-based discovery is accounting for modulation of protein-ligand interactions by ordered and bulk solvent. To investigate this, we compared ligand binding to a buried cavity in Cytochrome c Peroxidase (CcP), where affinity is dom

Proteins fluctuate between alternative conformations, which presents a challenge for ligand discovery because such flexibility is difficult to treat computationally owing to problems with conformational sampling and energy weighting. Here we describe a fle

Cardiac output (CO) measurement is crucial for the guidance of therapeutic decisions in critically ill and high-risk surgical patients. Newly developed completely non-invasive CO technologies are commercially available; however, their accuracy and precision have not recently been evaluated in a meta-analysis. We conducted a systematic search using PubMed, Cochrane Library of Clinical Trials, Scopus, and Web of Science to review published data comparing CO measured by bolus thermodilution with commercially available non-invasive technologies including pulse wave transit time, non-invasive pulse contour analysis, thoracic electrical bioimpedance/bioreactance, and CO2 rebreathing. The non-invasive CO technology was considered acceptable if the pooled estimate of percentage error was <30%, as previously recommended. Using a random-effects model, sd, pooled mean bias, and mean percentage error were calculated. An I2 statistic was also used to evaluate the inter-study heterogeneity. A total of 37 studies (1543 patients) were included. Mean CO of both methods was 4.78 litres min−1. Bias was presented as the reference method minus the tested methods in 15 studies. Only six studies assessed the random error (repeatability) of the tested device. The overall random-effects pooled bias (limits of agreement) and the percentage error were −0,13 [−2.38 , 2.12] litres min−1 and 47%, respectively. Inter-study sensitivity heterogeneity was high (I2=83%, P<0.001). With a wide percentage error, completely non-invasive CO devices are not interchangeable with bolus thermodilution. Additional studies are warranted to demonstrate their role in improving the quality of care.

 Characterizing flow patterns and mixing of fossil fuel-derived CO₂ is important for effectively using atmospheric measurements to constrain emissions inventories. Here we used measurements and a model of atmospheric radiocarbon (¹⁴C) to investigate the distribution and fluxes of atmospheric fossil fuel CO₂ across the state of California. We sampled ¹⁴C in annual C₃ grasses at 128 sites and used these measurements to test a regional model that simulated anthropogenic and ecosystem CO₂ fluxes, transport in the atmosphere, and the resulting Δ¹⁴C of annual grasses (Δ _g ). Average measured Δ _g levels in Los Angeles, San Francisco, the Central Valley, and the North Coast were 27.7 ± 20.0, 44.0 ± 10.9, 48.7 ± 1.9, and 59.9 ± 2.5‰, respectively, during the 2004–2005 growing season. Model predictions reproduced regional patterns reasonably well, with estimates of 27.6 ± 2.4, 39.4 ± 3.9, 46.8 ± 3.0, and 59.3 ± 0.2‰ for these same regions and corresponding to fossil fuel CO₂ mixing ratios (C _f ) of 13.7, 6.1, 4.8, and 0.3 ppm. Δ _g spatial heterogeneity in Los Angeles and San Francisco was higher in the measurements than in the predictions, probably from insufficient spatial resolution in the fossil fuel inventories (e.g., freeways are not explicitly included) and transport (e.g., within valleys). We used the model to predict monthly and annual transport patterns of fossil fuel-derived CO₂ within and out of California. Fossil fuel CO₂ emitted in Los Angeles and San Francisco was predicted to move into the Central Valley, raising C _f above that expected from local emissions alone. Annually, about 21, 39, 35, and 5% of fossil fuel emissions leave the California airspace to the north, east, south, and west, respectively, with large seasonal variations in the proportions. Positive correlations between westward fluxes and Santa Ana wind conditions were observed. The southward fluxes over the Pacific Ocean were maintained in a relatively coherent flow within the marine boundary layer, while the eastward fluxes were more vertically dispersed. Our results indicate that state and continental scale atmospheric inversions need to consider areas where mixing ratio measurements are sparse (e.g., over the ocean to the south and west of California), transport within and across the marine boundary layer, and terrestrial boundary layer dynamics. Radiocarbon measurements can be very useful in constraining these estimates.

OBJECTIVE: To improve the understanding of the natural disease progression of retinitis pigmentosa GTPase regulator (RPGR)-associated X-linked retinitis pigmentosa (XLRP). DESIGN: A multicenter, prospective, observational natural history study over 24 months. PARTICIPANTS: Male participants aged ≥7 years with a pathogenic variant in the RPGR gene, a best-corrected visual acuity (BCVA) score of ≥34 ETDRS letters, and a mean 68-loci retinal sensitivity (assessed by microperimetry) of 0.1 to 20 decibels (dB). METHODS: Participants were divided into subgroups based on their BCVA score at baseline: 34 to 73 (lower BCVA) or ≥74 (higher BCVA) ETDRS letters. There were 7 visits over 24 months. MAIN OUTCOME MEASURES: Change from baseline in BCVA, retinal sensitivity, low luminance visual acuity (LLVA), fixation stability, contrast sensitivity, visual field, anatomical measures, 25-item Visual Function Questionnaire (VFQ-25), intraocular pressure, and adverse events (AEs). RESULTS: Overall, 201 participants were included. The mean (standard deviation [SD]) age was 30.3 (11.9) years in the lower BCVA subgroup (n = 170) and 27.7 (10.1) years in the higher BCVA subgroup (n = 31). The study eye baseline mean (SD) BCVA scores were 59.4 (10.30) and 77.3 (3.95) in the lower and higher BCVA subgroups, respectively; the lower BCVA subgroup had lower retinal sensitivity in the study eye at baseline than the higher BCVA subgroup. Over 24 months, there were small observed changes in BCVA, retinal sensitivity, LLVA, fixation, contrast sensitivity, and fundus photography findings. There were observed mean (SD) changes at 24 months in the lower and higher BCVA subgroups of -1.01 (4.67) and 0.03 (5.83) dB-steradians in the volume of full-field hill of vision, -330.6 (869.51) and -122.7 (22.01) μm in distance from foveal center to the nearest border of preserved fundus autofluorescence, -104.3 (277.80) and -207.1 (171.01) μm in central ellipsoid width, and -2.8 (9.7) and -0.6 (7.6) in VFQ-25 composite score, respectively. There was 1 death from completed suicide. There were no ocular serious adverse events, and most AEs were mild/moderate. CONCLUSIONS: This study provides evidence of the slow natural progression of XLRP over 24 months in both subgroups and provides important functional, anatomical, and safety data. FINANCIAL DISCLOSURES: Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.