Browsing by Author "Tao, Jane"

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    Modeling protein structural ensembles using AWSEM-Suite
    (2023-04-11) Jin, Shikai; Tao, Jane; Wolynes, Peter
    Proteins are the driving force behind most cellular processes. Traditional methods for determining protein structure are limited to probing only a few static structures of a given protein. However, molecular dynamics simulation presently allows for the determination of the dynamics of a protein. In this thesis, I introduce AWSEM-Suite, a coarse-grained force field that has recently shown good performance in protein structure prediction experiments. This force field is based on physical principles and neural network-based machine learning. I describe the composition of the force field, as well as two new energy terms: the template-based and coevolutionary-guided terms. After discussing the problem of protein structure prediction, I showcase how we built an online server for AWSEM-Suite, describing the path between the input and output. Additionally, I discuss other techniques for structural analysis, including frustration protein structure refinement, a physics-based method that complements current methods. In the last part of this thesis, I introduce two applications: solving phase problems in X-ray crystallography and exploring the translocation mechanism of bacteriophage T7 helicase gp4 using AWSEM-Suite. Our results show that AWSEM-Suite provides better results than the state-of-the-art program I-TASSER-MR in finding phase information for a given protein. Furthermore, AWSEM-Suite can successfully predict the key loops that interact with ssDNA during gp4 translocation. We explore the intermediate structures of action, highlighting the possible mechanism of the role of electrostatic effects exerted by the binding and release of ATP molecules. In these chapters, we explore several related questions concerning protein structure prediction, protein structure refinement, and specific biological questions using AWSEM-Suite. In summary, our studies demonstrate that AWSEM-Suite is a powerful technology for exploring protein dynamics and predicting protein structure.
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    Using C. elegans to Identify Novel Targets Against Multidrug-Resistant Bacteria
    (2020-07-06) Hummell, Nicholas Andrew; Kirienko, Natasha; Tao, Jane
    Antibiotic-resistant infections cause an average of 23,000 deaths per year. Due to dwindling commercial interest for antimicrobial discovery, novel methods for combating infection and developing new antimicrobials are desperately needed. Previously in our lab, we performed a high-throughput chemical screen to identify small molecules that rescued the nematode Caenorhabditis elegans from infection by P. aeruginosa. Of the hits identified, 5 (LK32, LK34, LK35, LK38 and LK56) were determined to be stimulators of host defense pathways while 5 additional hits (DMAQ-B1, CD437, carboplatin, oxaliplatin, and PSB-069) possessed a known target or bioactivity but had no previously reported antimicrobial activity against P. aeruginosa.. Using microarray analysis, RNAi knockdown of candidate pathways, transgene reporters, and infection assays with other pathogens, we made important observations concerning the mechanism of action and therapeutic repurposing potential for the compounds. Firstly, I identified a subunit of the Mediator complex, mdt-15, and the PMK-1/p38 MAPK pathway as necessary for rescue for LK56 and LK38 respectively, demonstrating that both pathways are amenable to immune stimulation. I have also found that some molecules can defend against E. faecalis and S. aureus as well as Pseudomonas while being largely non-toxic. Additionally, most molecules stimulated the activation of multiple innate immune pathways. These experiments showed the potential for development of broad-spectrum immune stimulants and identified promising pathways amenable to immune stimulation. They also highlighted potential for our LK molecules as tools for future studies of innate immune stimulation in C. elegans. For our bioactive compounds, I used similar C. elegans-based methods to generate a number of important conclusions: I confirmed the antimicrobial activity of CD437 against Gram-positive pathogens, observed a weakness of P. aeruginosa to platinum complexes, and established the naturally occurring insulin mimetic, DMAQ-B1, as a powerful antimicrobial agent. Although toxic, an existing non-toxic analog presents potential for further therapeutic optimization. Through these studies, I have utilized C. elegans as a powerful drug discovery tool to gain insight into mechanism and therapeutic utility of two groups of anti-infective molecules. I have shown the strength of our model in drug repurposing efforts as well as demonstrated therapeutic potential for immune stimulation as a promising approach to combatting the growing antimicrobial resistance crisis.