Patel, Trushar
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- ItemProceedings of the 25th Analytical Ultracentrifugation workshops and symposium(Springer, 2023) Demeler, Borries; Gilbert, Robert; Patel, Trushar R.The 25th International Analytical Ultracentrifugation (AUC) Workshops and Symposium (AUC2022) took place at the University of Lethbridge in Lethbridge, Canada, in July 2022. In total, 104 attendees (Attendance Profile: 104 attendees, 69 in-person, 35 remote. Brazil 1, Canada 24, China 1, Czech Republic 2, Finland 1, France 3, Germany 22, India 3, Italy 1, Japan 4, Spain 1, Switzerland 3, Taiwan 1, United Kingdom 5, United States 32) participated in the event and presented the latest advances in the field. While the primary focus of the conference was to showcase the applications of AUC in chemical, life sciences, and nanoparticle disciplines, several presentations also integrated complementary methods, such as isothermal titration calorimetry, microscale thermophoresis, light scattering (static and dynamic), small-angle X-ray scattering, X-ray crystallography, and cryo-electron microscopy. Additionally, the delegates gained valuable hands-on experience from 20 workshops covering a broad range of applications, experimental designs and systems, and the latest software innovations in solution biophysics. The AUC2022 special volume highlights the sustained innovation, utility and relevance of AUC and related solution biophysical methods across various disciplines, including biochemistry, structural biology, synthetic polymer chemistry, carbohydrate chemistry, protein and nucleic acid characterization, nano-science, and macromolecular interactions.
- ItemCrystal structure of the TreS:Pep2 complex, initiating α-glucan synthesis in the GlgE pathway of mycobacteria(American Society for Biochemistry and Molecular Biology, 2019) Kermani, Ali A.; Roy, Rana; Gopalasingam, Chai; Kocurek, Klaudia I.; Patel, Trushar R.; Alderwick, Luke J.; Besra, Gurdyal S.; Fütterer, KlausA growing body of evidence implicates the mycobacterial capsule, the outermost layer of the mycobacterial cell envelope, in modulation of the host immune response and virulence of mycobacteria. Mycobacteria synthesize the dominant capsule component, α(1→4)-linked glucan, via three interconnected and potentially redundant metabolic pathways. Here, we report the crystal structure of the Mycobacterium smegmatis TreS:Pep2 complex, containing trehalose synthase (TreS) and maltokinase (Pep2), which converts trehalose to maltose 1-phosphate as part of the TreS:Pep2–GlgE pathway. The structure, at 3.6 Å resolution, revealed that a diamond-shaped TreS tetramer forms the core of the complex and that pairs of Pep2 monomers bind to opposite apices of the tetramer in a 4 + 4 configuration. However, for the M. smegmatis orthologues, results from isothermal titration calorimetry and analytical ultracentrifugation experiments indicated that the prevalent stoichiometry in solution is 4 TreS + 2 Pep2 protomers. The observed discrepancy between the crystallized complex and the behavior in the solution state may be explained by the relatively weak affinity of Pep2 for TreS (Kd 3.5 μm at mildly acidic pH) and crystal packing favoring the 4 + 4 complex. Proximity of the ATP-binding site in Pep2 to the complex interface provides a rational basis for rate enhancement of Pep2 upon binding to TreS, but the complex structure appears to rule out substrate channeling between the active sites of TreS and Pep2. Our findings provide a structural model for the trehalose synthase:maltokinase complex in M. smegmatis that offers critical insights into capsule assembly.
- ItemCanadian science meets parliament: building relationships between scientists and policymakers(Oxford University Press, 2021) Zhao, Jiaying; Azad, Meghan B.; Bertrand, Erin M.; Burton, Cole; Crooks, Valorie A.; Dawson, Jackie; Ford, Adam T.; Kaida, Angela; Krishnaswamy, Arjun; Kuok, Chikin; Mah, Catherine L.; McTaggart, Matt; Moehring, Amanda J.; Robert, Dominique; Schulte-Hostedde, Albrecht; Sparling, Heather; De Vera, Mary A.; Waterman, Stephanie; Patel, Trushar R.The first Science Meets Parliament event in Canada was held in November 2018 in Ottawa, where twenty-eight Tier II Canada Research Chairs (a specific class of Canadian university professor acknowledged by their peers as having the potential to lead in their field) from diverse disciplines met with forty-three Members of Canadian Parliament and Senators. The main goal of this event was to facilitate communication between these two key pillars of the society, to promote mutual understanding of the nature of their respective work, roles, and responsibilities, and to build long-term relationships. Here, we, representatives of the first cohort of scientists to participate in the program, summarize our experiences and lessons learned from this event, as well as our assessment of the benefits of attending this event for scientists, policy decision-makers, and institutions. Furthermore, we provide suggestions for similar future events in Canada and elsewhere.
- ItemInvestigating RNA-RNA interactions through computational and biophysical analysis(Oxford University Press, 2023) Mrozowich, Tyler; Park, Sean M.; Waldl, Maria; Henrickson, Amy; Tersteeg, Scott; Nelson, Corey R.; De Klerk, Anneke; Demeler, Borries; Hofacker, Ivo L.; Wolfinger, MIchael T.; Patel, Trushar R.Numerous viruses utilize essential long-range RNA–RNA genome interactions, specifically flaviviruses. Using Japanese encephalitis virus (JEV) as a model system, we computationally predicted and then biophysically validated and characterized its long-range RNA–RNA genomic interaction. Using multiple RNA computation assessment programs, we determine the primary RNA–RNA interacting site among JEV isolates and numerous related viruses. Following in vitro transcription of RNA, we provide, for the first time, characterization of an RNA–RNA interaction using size-exclusion chromatography coupled with multi-angle light scattering and analytical ultracentrifugation. Next, we demonstrate that the 5′ and 3′ terminal regions of JEV interact with nM affinity using microscale thermophoresis, and this affinity is significantly reduced when the conserved cyclization sequence is not present. Furthermore, we perform computational kinetic analyses validating the cyclization sequence as the primary driver of this RNA–RNA interaction. Finally, we examined the 3D structure of the interaction using small-angle X-ray scattering, revealing a flexible yet stable interaction. This pathway can be adapted and utilized to study various viral and human long-non-coding RNA–RNA interactions and determine their binding affinities, a critical pharmacological property of designing potential therapeutics.
- ItemThe biomedical and bioengineering potential of protein nanocompartments(Elsevier, 2020) Demchuk, Aubrey M.; Patel, Trushar R.Protein nanocompartments (PNCs) are self-assembling biological nanocages that can be harnessed as platforms for a wide range of nanobiotechnology applications. The most widely studied examples of PNCs include virus-like particles, bacterial microcompartments, encapsulin nanocompartments, enzyme-derived nanocages (such as lumazine synthase and the E2 component of the pyruvate dehydrogenase complex), ferritins and ferritin homologues, small heat shock proteins, and vault ribonucleoproteins. Structural PNC shell proteins are stable, biocompatible, and tolerant of both interior and exterior chemical or genetic functionalization for use as vaccines, therapeutic delivery vehicles, medical imaging aids, bioreactors, biological control agents, emulsion stabilizers, or scaffolds for biomimetic materials synthesis. This review provides an overview of the recent biomedical and bioengineering advances achieved with PNCs with a particular focus on recombinant PNC derivatives.