Engineered Toxins “Zymoxins” Are Activated by the HCV NS3 Protease by Removal of an Inhibitory Protein Domain

The synthesis of inactive enzyme precursors, also known as “zymogens,” serves as a mechanism for regulating the execution of selected catalytic activities in a desirable time and/or site. Zymogens are usually activated by proteolytic cleavage. Many viruses encode proteases that execute key proteolytic steps of the viral life cycle. Here, we describe a proof of concept for a therapeutic approach to fighting viral infections through eradication of virally infected cells exclusively, thus limiting virus production and spread. Using the hepatitis C virus (HCV) as a model, we designed two HCV NS3 protease-activated “zymogenized” chimeric toxins (which we denote “zymoxins”). In these recombinant constructs, the bacterial and plant toxins diphtheria toxin A (DTA) and Ricin A chain (RTA), respectively, were fused to rationally designed inhibitor peptides/domains via an HCV NS3 protease-cleavable linker. The above toxins were then fused to the binding and translocation domains of Pseudomonas exotoxin A in order to enable translocation into the mammalian cells cytoplasm. We show that these toxins exhibit NS3 cleavage dependent increase in enzymatic activity upon NS3 protease cleavage in vitro. Moreover, a higher level of cytotoxicity was observed when zymoxins were applied to NS3 expressing cells or to HCV infected cells, demonstrating a potential therapeutic window. The increase in toxin activity correlated with NS3 protease activity in the treated cells, thus the therapeutic window was larger in cells expressing recombinant NS3 than in HCV infected cells. This suggests that the “zymoxin” approach may be most appropriate for application to life-threatening acute infections where much higher levels of the activating protease would be expected

Benchmarking of Data-Driven Causality Discovery Approaches in the Interactions of Arctic Sea Ice and Atmosphere

The Arctic sea ice has retreated rapidly in the past few decades, which is believed to be driven by various dynamic and thermodynamic processes in the atmosphere. The newly open water resulted from sea ice decline in turn exerts large influence on the atmosphere. Therefore, this study aims to investigate the causality between multiple atmospheric processes and sea ice variations using three distinct data-driven causality approaches that have been proposed recently: Temporal Causality Discovery Framework Non-combinatorial Optimization via Trace Exponential and Augmented lagrangian for Structure learning (NOTEARS) and Directed Acyclic Graph-Graph Neural Networks (DAG-GNN). We apply these three algorithms to 39 years of historical time-series data sets, which include 11 atmospheric variables from ERA-5 reanalysis product and passive microwave satellite retrieved sea ice extent. By comparing the causality graph results of these approaches with what we summarized from the literature, it shows that the static graphs produced by NOTEARS and DAG-GNN are relatively reasonable. The results from NOTEARS indicate that relative humidity and precipitation dominate sea ice changes among all variables, while the results from DAG-GNN suggest that the horizontal and meridional wind are more important for driving sea ice variations. However, both approaches produce some unrealistic cause-effect relationships. Additionally, these three methods cannot well detect the delayed impact of one variable on another in the Arctic. It also turns out that the results are rather sensitive to the choice of hyperparameters of the three methods. As a pioneer study, this work paves the way to disentangle the complex causal relationships in the Earth system, by taking the advantage of cutting-edge Artificial Intelligence technologies. © Copyright © 2021 Huang, Kleindessner, Munishkin, Varshney, Guo and Wang.Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Ribosome-induced changes in elongation factor Tu conformation control GTP hydrolysis

In translation, elongation factor Tu (EF-Tu) molecules deliver aminoacyl-tRNAs to the mRNA-programmed ribosome. The GTPase activity of EF-Tu is triggered by ribosome-induced conformational changes of the factor that play a pivotal role in the selection of the cognate aminoacyl-tRNAs. We present a 6.7-Å cryo-electron microscopy map of the aminoacyl-tRNA·EF-Tu·GDP·kirromycin-bound Escherichia coli ribosome, together with an atomic model of the complex obtained through molecular dynamics flexible fitting. The model reveals the conformational changes in the conserved GTPase switch regions of EF-Tu that trigger hydrolysis of GTP, along with key interactions, including those between the sarcin-ricin loop and the P loop of EF-Tu, and between the effector loop of EF-Tu and a conserved region of the 16S rRNA. Our data suggest that GTP hydrolysis on EF-Tu is controlled through a hydrophobic gate mechanism