Structure and function of CutC choline lyase from human microbiota bacterium Klebsiella pneumoniae

Publisher Copyright: © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.CutC choline trimethylamine-lyase is an anaerobic bacterial glycyl radical enzyme (GRE) that cleaves choline to produce trimethylamine (TMA) and acetaldehyde. In humans, TMA is produced exclusively by the intestinal microbiota, and its metabolite, trimethylamine oxide, has been associated with a higher risk of cardiovascular diseases. Therefore, information about the three-dimensional structures of TMA-producing enzymes is important for microbiota-targeted drug discovery. We have cloned, expressed, and purified the CutC GRE and the activating enzyme CutD from Klebsiella pneumoniae, a representative of the human microbiota. We have determined the first crystal structures of both the choline-bound and choline-free forms of CutC and have discovered that binding of choline at the ligand-binding site triggers conformational changes in the enzyme structure, a feature that has not been observed for any other characterized GRE.publishersversionPeer reviewe

Robustness of empirical vibration correlation techniques for predicting the instability of unstiffened cylindrical composite shells in axial compression

Thin-walled carbon fiber reinforced plastic (CFRP) shells are increasingly used in aerospace industry. Such shells are prone to the loss of stability under compressive loads. Furthermore, the instability onset of monocoque shells exhibits a pronounced imperfection sensitivity. The vibration correlation technique (VCT) is being developed as a nondestructive test method for evaluation of the buckling load of the shells. In this study, accuracy and robustness of an existing and a modified VCT method are evaluated. With this aim, more than 20 thin-walled unstiffened CFRP shells have been produced and tested. The results obtained suggest that the vibration response under loads exceeding 0.25 of the linear buckling load needs to be characterized for a successful application of the VCT. Then the largest unconservative discrepancy of prediction by the modified VCT method amounted to ca. 22% of the critical load. Applying loads exceeding 0.9 of the buckling load reduced the average relative discrepancy to 6.4%.</p

Robustness of empirical vibration correlation techniques for predicting the instability of unstiffened cylindrical composite shells in axial compression

Thin-walled carbon fiber reinforced plastic (CFRP) shells are increasingly used in aerospace industry. Such shells are prone to the loss of stability under compressive loads. Furthermore, the instability onset of monocoque shells exhibits a pronounced imperfection sensitivity. The vibration correlation technique (VCT) is being developed as a nondestructive test method for evaluation of the buckling load of the shells. In this study, accuracy and robustness of an existing and a modified VCT method are evaluated. With this aim, more than 20 thin-walled unstiffened CFRP shells have been produced and tested. The results obtained suggest that the vibration response under loads exceeding 0.25 of the linear buckling load needs to be characterized for a successful application of the VCT. Then the largest unconservative discrepancy of prediction by the modified VCT method amounted to ca. 22% of the critical load. Applying loads exceeding 0.9 of the buckling load reduced the average relative discrepancy to 6.4%.Aerospace Structures & Computational Mechanic

Discovery of SARS-CoV-2 Nsp14 and Nsp16 Methyltransferase Inhibitors by High-Throughput Virtual Screening

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) uses mRNA capping to evade the human immune system. The cap formation is performed by the SARS-CoV-2 mRNA cap methyltransferases (MTases) nsp14 and nsp16, which are emerging targets for the development of broad-spectrum antiviral agents. Here, we report results from high-throughput virtual screening against these two enzymes. The docking of seven million commercially available drug-like compounds and S-adenosylmethionine (SAM) co-substrate analogues against both MTases resulted in 80 virtual screening hits (39 against nsp14 and 41 against nsp16), which were purchased and tested using an enzymatic homogeneous time-resolved fluorescent energy transfer (HTRF) assay. Nine compounds showed micromolar inhibition activity (IC50 &lt; 200 &mu;M). The selectivity of the identified inhibitors was evaluated by cross-checking their activity against human glycine N-methyltransferase. The majority of the compounds showed poor selectivity for a specific MTase, no cytotoxic effects, and rather poor cell permeability. Nevertheless, the identified compounds represent good starting points that have the potential to be developed into efficient viral MTase inhibitors