Browsing by Author "Wang, Xiaoling"

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    MRI and NMR Investigations of Transport in Soft Materials and Explorations of Electron-Nuclear Interactions for Liquid-State Dynamic Nuclear Polarization
    Wang, Xiaoling (Virginia Tech, 2015-08-28)
    The first part of this dissertation (Chapters 1 to 4) describes the use of magnetic resonance techniques for polymeric material characterizations in solutions, with emphasis on methods utilizing magnetic field gradients - magnetic resonance imaging (MRI) and pulsed-field-gradient (PFG) NMR. The second part (Chapter 5) presents enhancements to dynamic nuclear polarization, an intensity enhancement approach for magnetic resonance techniques. In Chapter 2, I illustrate a characterization method to quantify free polymer chain content in a polymer/DNA complex (polyplex) formulation via one-dimensional proton NMR experiments. This assessment of free polymer quantity has critical impacts on in vivo gene transfection efficiency, cellular uptake, as well as toxicity of polycationic gene delivery vectors. Specifically, I investigated the complexation properties of three different polymeric "theranostic" agents, which combine an imaging functionality on the polymer as well as a DNA/RNA complexation component. These agents are under development to allow real time clinical monitoring of drug delivery and efficacy using MRI. Our NMR method provides simple and quantitative assessment of free and DNA-complexed polymers, including the actual polymer amine to DNA phosphate molar ratio (N/P ratio) within polyplexes. The NMR results are in close agreement with the stoichiometric number of polymer/DNA binding obtained by isothermal titration calorimetry. The noninvasive nature of this method allows broad application to a range of polyelectrolyte coacervates, for understanding and optimizing polyelectrolyte complex formation. Chapter 3 demonstrates a time-resolved MRI approach for measuring diffusion of drug-delivery polymeric nanoparticles on mm to cm scales as well as monitoring nanoparticle concentration distribution in bulk biological hydrogels. Our results show that as the particle size and surface charge become larger, collagen gel at tumor relevant concentration (1.0 wt.%) presents a more significant impediment to the diffusive transport of negatively charged nanoparticles. These results agree well with those obtained by fluorescence spectroscopies (neutral or slightly positively charged diffusing particles) as well as the proposed electrostatic bandpass theory of tumor interstitium (negatively charged particles). This study provides fundamental information for the design of polymeric theranostic vectors and carries implications that would benefit the understanding of nanoparticle transport in solid tumors. Furthermore, this work takes a significant step toward developing quantitative and real time in vivo monitoring of clinical drug delivery using MRI. Chapter 4 addresses the application of PFG-NMR for the determination of weight-average molar mass (Mw) for polyanions that have anti-HIV activity through the measurement of polymer diffusion coefficients in solutions. The effective characterization of molecular weights of polyelectrolytes has been a general and growing problem for the polymer industry, with no clear solutions in sight. In this study, we obtained the molar masses (Mw) for two series of sulfonated copolymers using sodium polystyrene sulfonate samples as molecular weight standards. PFG-NMR has notable advantages over conventional techniques for the characterization of charged polymers and shows great promise for becoming an effective alternative to chromatography methods. Chapter 5 is devoted to experimental and theoretical studies of liquid state dynamic nuclear polarization (DNP) via the Overhauser effect. Based on the adventurous work done by previous Dorn group members, we show that for 1H-nuclide-containing systems, the dipolar DNP enhancement can be significantly improved by decreasing the correlation time of the interaction by utilizing a supercritical fluid (SF CO2) which allows for greater dipolar enhancements at higher magnetic fields. For molecules containing the ubiquitous 13C nuclide, we show that previously unreported sp hybridized (H-C) alkyne systems represented by the phenylacetylene-nitroxide system exhibit very large scalar-dominated enhancements. Furthermore, we show for a wide range of molecular systems that the Fermi contact interaction can be computationally predicted via electron-nuclear hyperfine coupling and correlated with experimental 13C DNP enhancements. For biomedical applications, the enhancement of metabolites in SF CO2 followed by rapid dissolution in water or biological fluids is an attractive approach for future hyperpolarized NMR and MRI applications. Moreover, with the aid of density functional theory calculations, solution state DNP provides a unique approach for studying intermolecular weak bonding interaction of solutes in normal liquids and SF fluids.
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    RETICULON-LIKE PROTEIN B2 is a proviral factor co-opted for the biogenesis of viral replication organelles in plants
    Zhang, Qianshen; Wen, Zhiyan; Zhang, Xin; She, Jiajie; Wang, Xiaoling; Gao, Zongyu; Wang, Ruiqi; Zhao, Xiaofei; Su, Zhen; Li, Zhen; Li, Dawei; Wang, Xiaofeng; Zhang, Yongliang (Oxford University Press, 2023-05)
    Endomembrane remodeling to form a viral replication complex (VRC) is crucial for a virus to establish infection in a host. Although the composition and function of VRCs have been intensively studied, host factors involved in the assembly of VRCs for plant RNA viruses have not been fully explored. TurboID-based proximity labeling (PL) has emerged as a robust tool for probing molecular interactions in planta. However, few studies have employed the TurboID-based PL technique for investigating plant virus replication. Here, we used Beet black scorch virus (BBSV), an endoplasmic reticulum (ER)-replicating virus, as a model and systematically investigated the composition of BBSV VRCs in Nicotiana benthamiana by fusing the TurboID enzyme to viral replication protein p23. Among the 185 identified p23-proximal proteins, the reticulon family of proteins showed high reproducibility in the mass spectrometry data sets. We focused on RETICULON-LIKE PROTEIN B2 (RTNLB2) and demonstrated its proviral functions in BBSV replication. We showed that RTNLB2 binds to p23, induces ER membrane curvature, and constricts ER tubules to facilitate the assembly of BBSV VRCs. Our comprehensive proximal interactome analysis of BBSV VRCs provides a resource for understanding plant viral replication and offers additional insights into the formation of membrane scaffolds for viral RNA synthesis. TurboID-based proximity labeling reveals the viral replication complex structure of a plant virus, unveiling a proviral function of RETICULON-LIKE PROTEIN B2 in viral replication complex formation.