Browsing by Subject "Membrane Proteins"

Now showing 1 - 5 of 5
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    The Effect of α-Synuclein on Lipid Membrane Properties Characterized by Molecular Dynamics and Atomic Force Microscopy
    (2018-08) Brummel, Benjamin
    The protein α-synuclein (αSyn), primarily recognized for its link to neurodegenerative disorders, has multiple reported functions. One well-established role of αSyn is its ability to bind and remodel lipid membranes. This ability has been characterized in synthetic lipid bilayers and has been observed both in cellular and in vivo models. The native environment of αSyn—the presynaptic terminal of neurons—contains mitochondria and synaptic vesicles, which have unique membranes that differ from previously studied models. The goal of this dissertation was to characterize how lipids enriched in synaptic vesicles and mitochondria affect how αSyn changes membrane properties. First, molecular dynamics (MD) simulations of synaptic vesicle-mimic bilayers showed how lipids with polyunsaturated fatty acids modify membrane properties and interact with αSyn. Next, tubulation experiments were combined with MD simulations to explore how αSyn remodels bilayers containing cardiolipin and phosphatidylethanolamine, two lipids enriched in mitochondria. Finally, methods were developed to characterize lipid vesicle mechanical properties using pulsed force mode (PFM) atomic force microscopy (AFM). This work provides insight into the specifics of how αSyn affects the properties of synaptic vesicle and mitochondrial membranes and demonstrates how PFM-AFM can identify the mechanical properties of lipid vesicles.
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    Ensembles from dynamic refinement of non-phosphorylated and phosphorylated phospholamban-SERCA complexes
    (2021-01-22) Weber, Daniel K; Sanz-Hernández, Máximo; Uddigiri, Venkateswara Reddy; Wang, Songlin; Larsen, Erik K; Gopinath, Tata; Gustavsson, Martin; Cornea, Razvan L; Thomas, David D; De Simone, Alfonso; Veglia, Gianluigi; vegli001@umn.edu; Veglia, Gianluigi; Veglia Lab
    Analysis, setup and ensembles of the dynamic refinement of phospholamban-SERCA complexes in the calcium-free E2 state. Includes trajectory files, analytic scripts, restraint files, setup files, simulation code, raw data (including OS-ssNMR data) and processed data used for the associated citations as per transparent reporting requirements.
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    Molecular dynamics simulations of membranes and membrane proteins.
    (2011-07) Perlmutter, Jason David
    Membranes composed of a lipid bilayer and embedded proteins are ubiquitous in nature. They form the barrier which demarcates every cell from its environment and separates the distinct organelles within eukaryotic cells, implicating membranes in a wide range of biological processes. The function of membranes and membrane proteins are determined by their structure, and the central focus of this thesis is the use of computational molecular dynamics simulations to study experimentally inaccessible details of membrane structure. Firstly, we have simulated ternary lipid bilayers containing steroids with a range of headgroup hydrophobicities, observing a correlation between the membrane lateral organization and the orientation of the steroid. Based on these results we suggest a general framework to distinguish previously identified steroid domain promoters and inhibitors. Secondly, we investigate the role of interleaflet coupling in membrane structure. This includes describing a compositional dependence to the interleaflet organization of phase separated membranes, as well as investigating structural perturbations due to interleaflet differences in composition. Thirdly, we demonstrate a strategy for obtaining experimental verification through low angle X-ray scattering and discuss its potential application to complex phase separated mixtures. The second focus of this thesis is considering how the structural features of membranes affect the behavior of membrane proteins. The membrane protein α-Synuclein is of wide interest due to its association with Parkinson's Disease, but its physiological function remains unknown. A third focus of this thesis is the structure of membrane-mimetics, such as detergent micelles and amphipathic polymers, which are commonly used for the stabilization of membrane proteins. Their potential distinct influence on protein behavior currently remains an unresolved hindrance to experimental characterization. The simulations presented herein demonstrate a distinct effect of membrane curvature on α-Synuclein behavior and suggest a potential role in regulating vesicle fusion. Collectively, these simulations of model systems offer insight into the fundamental features which determine the behavior of complex biological membranes.
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    Replica-averaged orientational-restrained ensembles of DWORF and P15A-DWORF in lipid bilayers
    (2021-05-06) Uddigiri, Venkateswara Reddy; Weber, Daniel K; Wang, Songlin; Larsen, Erik K; Gopinath, Tata; De Simone, Alfonso; Robia, Seth; Veglia, Gianluigi; vegli001@umn.edu; Veglia, Gianluigi
    Replica-averaged orientational-restrained molecular dynamics (RAOR-MD) ensembles of DWORF and the DWORF-P15A mutant in lipid bilayers. Restrained to 15N chemical shift anisotropy and 1H-15N dipolar coupling data obtained from oriented sample solid-state NMR spectroscopy.
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    Structural Characterization of Sarcolipin by Solid State NMR and Investigation of its Role in the Regulation of Sarco(endo)plasmic Reticulum Calcium Adenosine-Triphospatase
    (2014-02) Mote, Kaustubh
    Structural characterization of membrane proteins and their complexes is an important and ever growing challenge to the classical techniques of biomolecular structural characterization. Rapid developments in the field of solid state NMR (ssNMR) have opened up an exciting new alternative to X-ray crystallography, as these studies can now be performed in fully hydrated lipid bilayers that faithfully mimic the physiologically relevant conditions. Nonetheless, routine application of ssNMR on biomolecular systems is hampered by their low sensitivity and spectral resolution. In this work, we have addressed these challenges by developing new strategies to study membrane proteins by ssNMR. With a set of improved pulse sequences for oriented and magic angle spinning techniques in ssNMR, we determined the topology (i.e. the structure and transmembrane orientation) of sarcolipin, a regulator of the Sarco(endo)plasmic Reticulum Ca2+-ATPase (SERCA), in lipid bilayers. These techniques are further used to study the complex between these two proteins and understand the molecular basis for this regulatory interaction. The methodological developments reported here are transferable to studies on other membrane proteins and they clear several roadblocks in the successful application of ssNMR for these challenging bio-molecular systems. Finally, we present how these studies have furthered our understanding of the regulation of muscle relaxation process by SERCA. These findings represent the first steps in designing new therapeutic approaches for cardiac and skeletal muscle disorders.