2'-O methylation of RNA cap in SARS-CoV-2 captured by serial crystallography.

The genome of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) coronavirus has a capping modification at the 5'-untranslated region (UTR) to prevent its degradation by host nucleases. These modifications are performed by the Nsp10/14 and Nsp10/16 heterodimers using S-adenosylmethionine as the methyl donor. Nsp10/16 heterodimer is responsible for the methylation at the ribose 2'-O position of the first nucleotide. To investigate the conformational changes of the complex during 2'-O methyltransferase activity, we used a fixed-target serial synchrotron crystallography method at room temperature. We determined crystal structures of Nsp10/16 with substrates and products that revealed the states before and after methylation, occurring within the crystals during the experiments. Here we report the crystal structure of Nsp10/16 in complex with Cap-1 analog (m7GpppAm2'-O). Inhibition of Nsp16 activity may reduce viral proliferation, making this protein an attractive drug target.

Fixed-target serial crystallography at the Structural Biology Center.

Serial synchrotron crystallography enables the study of protein structures under physiological temperature and reduced radiation damage by collection of data from thousands of crystals. The Structural Biology Center at Sector 19 of the Advanced Photon Source has implemented a fixed-target approach with a new 3D-printed mesh-holder optimized for sample handling. The holder immobilizes a crystal suspension or droplet emulsion on a nylon mesh, trapping and sealing a near-monolayer of crystals in its mother liquor between two thin Mylar films. Data can be rapidly collected in scan mode and analyzed in near real-time using piezoelectric linear stages assembled in an XYZ arrangement, controlled with a graphical user interface and analyzed using a high-performance computing pipeline. Here, the system was applied to two ß-lactamases: a class D serine ß-lactamase from Chitinophaga pinensis DSM 2588 and L1 metallo-ß-lactamase from Stenotrophomonas maltophilia K279a.

High-Throughput Virtual Screening and Validation of a SARS-CoV-2 Main Protease Noncovalent Inhibitor.

Despite the recent availability of vaccines against the acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the search for inhibitory therapeutic agents has assumed importance especially in the context of emerging new viral variants. In this paper, we describe the discovery of a novel noncovalent small-molecule inhibitor, MCULE-5948770040, that binds to and inhibits the SARS-Cov-2 main protease (Mpro) by employing a scalable high-throughput virtual screening (HTVS) framework and a targeted compound library of over 6.5 million molecules that could be readily ordered and purchased. Our HTVS framework leverages the U.S. supercomputing infrastructure achieving nearly 91% resource utilization and nearly 126 million docking calculations per hour. Downstream biochemical assays validate this Mpro inhibitor with an inhibition constant (Ki) of 2.9 µM (95% CI 2.2, 4.0). Furthermore, using room-temperature X-ray crystallography, we show that MCULE-5948770040 binds to a cleft in the primary binding site of Mpro forming stable hydrogen bond and hydrophobic interactions. We then used multiple µs-time scale molecular dynamics (MD) simulations and machine learning (ML) techniques to elucidate how the bound ligand alters the conformational states accessed by Mpro, involving motions both proximal and distal to the binding site. Together, our results demonstrate how MCULE-5948770040 inhibits Mpro and offers a springboard for further therapeutic design.

Increasing the bioavailability of Ru(III) anticancer complexes through hydrophobic albumin interactions.

A series of pyridine-based derivatives of the clinically successful Ru(III)-based complexes indazolium [trans-RuCl4(1H-indazole)2] (KP1019) and sodium [trans-RuCl4(1H-indazole)2] (KP1339) have been synthesized to probe the effect of hydrophobic interactions with human serum albumin (hsA) on anticancer activity. The solution behavior and protein interactions of the new compounds were characterized by using electron paramagnetic resonance (EPR) and UV/Vis spectroscopy. These studies have revealed that incorporation of hydrophobic substituents at the 4'-position of the axial pyridine ligand stabilizes non-coordinate interactions with hsA. As a consequence, direct coordination to the protein is inhibited, which is expected to increase the bioavailability of the complexes, thus potentially leading to improved anticancer activity. By using this approach, the lifetimes of hydrophobic protein interactions were extended from 2 h for the unsubstituted pyridine complex, to more than 24 h for several derivatives. Free complexes were tested for their anticancer activity against the SW480 human colon carcinoma cell line, exhibiting low cytotoxicity. Pre-treatment with hsA improved the solubility of every compound and led to some changes in activity. Particularly notable was the difference in activity between the methyl- and dibenzyl-functionalized complexes. The former shows reduced activity after incubation with hsA, indicating reduced bioavailability due to protein coordination. The latter exhibits little activity on its own but, following treatment with hsA, exhibited significant cytotoxicity, which is consistent with its ability to form non-coordinate interactions with the protein. Overall, our studies demonstrate that non-coordinate interactions with hsA are a viable target for enhancing the activity of Ru(III)-based complexes in vivo.

Deep learning at the edge enables real-time streaming ptychographic imaging.

Coherent imaging techniques provide an unparalleled multi-scale view of materials across scientific and technological fields, from structural materials to quantum devices, from integrated circuits to biological cells. Driven by the construction of brighter sources and high-rate detectors, coherent imaging methods like ptychography are poised to revolutionize nanoscale materials characterization. However, these advancements are accompanied by significant increase in data and compute needs, which precludes real-time imaging, feedback and decision-making capabilities with conventional approaches. Here, we demonstrate a workflow that leverages artificial intelligence at the edge and high-performance computing to enable real-time inversion on X-ray ptychography data streamed directly from a detector at up to 2 kHz. The proposed AI-enabled workflow eliminates the oversampling constraints, allowing low-dose imaging using orders of magnitude less data than required by traditional methods.

Pyridine analogues of the antimetastatic Ru(III) complex NAMI-A targeting non-covalent interactions with albumin.

A series of pyridine-based derivatives of the antimetastatic Ru(III) complex imidazolium [trans-RuCl(4)(1H-imidazole)(DMSO-S)] (NAMI-A) have been synthesized along with their sodium-ion compensated analogues. These compounds have been characterized by X-ray crystallography, electron paramagnetic resonance (EPR), NMR, and electrochemistry, with the goal of probing their noncovalent interactions with human serum albumin (hsA). EPR studies show that the choice of imidazolium ligands and compensating ions does not strongly influence the rates of ligand exchange processes in aqueous buffer solutions. By contrast, the rate of formation and persistence of interactions of the complexes with hsA is found to be strongly dependent on the properties of the axial ligands. The stability of noncovalent binding is shown to correlate with the anticipated ability of the various pyridine ligands to interact with the hydrophobic binding domains of hsA. These interactions prevent the oligomerization of the complexes in solution and limit the rate of covalent binding to albumin amino acid side chains. Electrochemical studies demonstrate relatively high reduction potentials for these complexes, leading to the formation of Ru(II) species in aqueous solutions containing biological reducing agents, such as ascorbate. However, EPR measurements indicate that while noncovalent interactions with hsA do not prevent reduction, covalent binding produces persistent mononuclear Ru(III) species under these conditions.

FAIR principles for AI models with a practical application for accelerated high energy diffraction microscopy.

A concise and measurable set of FAIR (Findable, Accessible, Interoperable and Reusable) principles for scientific data is transforming the state-of-practice for data management and stewardship, supporting and enabling discovery and innovation. Learning from this initiative, and acknowledging the impact of artificial intelligence (AI) in the practice of science and engineering, we introduce a set of practical, concise, and measurable FAIR principles for AI models. We showcase how to create and share FAIR data and AI models within a unified computational framework combining the following elements: the Advanced Photon Source at Argonne National Laboratory, the Materials Data Facility, the Data and Learning Hub for Science, and funcX, and the Argonne Leadership Computing Facility (ALCF), in particular the ThetaGPU supercomputer and the SambaNova DataScale® system at the ALCF AI Testbed. We describe how this domain-agnostic computational framework may be harnessed to enable autonomous AI-driven discovery.

Linking scientific instruments and computation: Patterns, technologies, and experiences.

Powerful detectors at modern experimental facilities routinely collect data at multiple GB/s. Online analysis methods are needed to enable the collection of only interesting subsets of such massive data streams, such as by explicitly discarding some data elements or by directing instruments to relevant areas of experimental space. Thus, methods are required for configuring and running distributed computing pipelines-what we call flows-that link instruments, computers (e.g., for analysis, simulation, artificial intelligence [AI] model training), edge computing (e.g., for analysis), data stores, metadata catalogs, and high-speed networks. We review common patterns associated with such flows and describe methods for instantiating these patterns. We present experiences with the application of these methods to the processing of data from five different scientific instruments, each of which engages powerful computers for data inversion,model training, or other purposes. We also discuss implications of such methods for operators and users of scientific facilities.

Canadian Plastic Surgery Resident Work Hour Restrictions: Practices and Perceptions of Residents and Program Directors.

BACKGROUND: The impact of resident work hour restrictions on training and patient care remains a highly controversial topic, and to date, there lacks a formal assessment as it pertains to Canadian plastic surgery residents. OBJECTIVE: To characterize the work hour profile of Canadian plastic surgery residents and assess the perspectives of residents and program directors regarding work hour restrictions related to surgical competency, resident wellness, and patient safety. METHODS: An anonymous online survey developed by the authors was sent to all Canadian plastic surgery residents and program directors. Basic summary statistics were calculated. RESULTS: Eighty (53%) residents and 10 (77%) program directors responded. Residents reported working an average of 73 hours in hospital per week with 8 call shifts per month and sleep 4.7 hours/night while on call. Most residents (88%) reported averaging 0 post-call days off per month and 61% will work post-call without any sleep. The majority want the option of working post-call (63%) and oppose an 80-hour weekly maximum (77%). Surgical and medical errors attributed to post-call fatigue were self-reported by 26% and 49% of residents, respectively. Residents and program directors expressed concern about the ability to master surgical skills without working post-call. CONCLUSIONS: The majority of respondents oppose duty hour restrictions. The reason is likely multifactorial, including the desire of residents to meet perceived expectations and to master their surgical skills while supervised. If duty hour restrictions are aggressively implemented, many respondents feel that an increased duration of training may be necessary.

HISTORIQUE: L'effet des restrictions des heures de travail des résidents sur la formation et les soins aux patients est un sujet très controversé. Jusqu'à présent, il n'y a pas d'évaluations officielles de cette réalité chez les résidents canadiens en chirurgie plastique. OBJECTIF: Caractériser le profil des heures de travail des résidents canadiens en chirurgie plastique et évaluer les points de vue des résidents et des directeurs de programme à l'égard de l'effet des restrictions des heures de travail sur la compétence chirurgicale, le bien-être des résidents et la sécurité des patients. MÉTHODOLOGIE: Les auteurs ont préparé un sondage anonyme en ligne qu'ils ont transmis à tous les résidents et les directeurs de programme en chirurgie plastique au Canada. Ils ont synthétisé les statistiques de base. RÉSULTATS: Au total, 80 résidents (53 %) et dix directeurs de programme (77 %) ont répondu au sondage. Les résidents ont déclaré faire une moyenne de 73 heures de travail hospitalier par semaine, faire huit quarts de garde par mois et dormir 4,7 heures par nuit lorsqu'ils sont sur appel. La plupart d'entre eux (88 %) déclarent une moyenne de 0 journée de congé après une garde, et 61 % travaillent ensuite sans avoir dormi. La majorité désire pouvoir travailler après une garde (63 %) et s'oppose à un maximum hebdomadaire de 80 heures (77 %). Par ailleurs, 26 % des résidents précisent avoir fait des erreurs chirurgicales et 49 %, des erreurs médicales qu'ils attribuent à la fatigue accumulée après une garde. Les résidents et les directeurs de programme s'inquiètent de la capacité des résidents à maîtriser les habiletés chirurgicales s'ils ne travaillent pas après les gardes. CONCLUSIONS: La majorité des répondants s'opposent aux restrictions des heures de garde. La raison est probablement multifactorielle, y compris le fait que les résidents souhaitent répondre aux attentes perçues et maîtriser leurs habiletés chirurgicales pendant qu'ils sont sous supervision. Si les restrictions des heures de garde étaient vigoureusement adoptées, de nombreux répondants croient qu'il faudrait allonger la formation.