Browsing by Subject "Nuclear magnetic resonance"

Now showing 1 - 3 of 3
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    Application of molecular modeling techniques to study the structure, dynamics, and interactions of membrane proteins.
    (2011-08) Shi, Lei
    Membrane proteins constitute ~30% of all the genomes and ~70% of the drug targets. However, less than 1% of the entries in the protein data bank are membrane proteins. The underrepresentation of membrane protein structures limits our understanding of their functions. This thesis summarizes my effects to apply theoretical methods to understand the structure and function relationships of membrane proteins. Specifically, we developed computational techniques to interpret solution and solid-state NMR data of membrane proteins and determine their high resolution structures. We further performed molecular dynamics simulations to study their dynamics, interaction with other proteins and the lipid bilayer environment. We applied these approaches to phospholamban, which is a membrane protein that is involved in cardiac muscle relaxation by regulating Ca2+-ATPase activity. Our results provide new insights to understand how membrane proteins elicit their function.
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    Characterization of the conformational states of phospholamban and their roles in regulation of SR Calcium-ATPase
    (2012-12) Gustavsson, Bengt Martin
    Membrane proteins constitute 30% of the human genome but represent only a small fraction of the known three-dimensional protein structures. In this thesis I describe the characterization of the membrane protein complex between sarcoplasmic reticulum Ca2+-ATPase (SERCA) and phospholamban (PLN). SERCA drives cardiac muscle relaxation by active transport of Ca2+ ions into the SR. PLN is a small membrane protein that consists of a helical trans-membrane domain connected to a cytoplasmic domain through a short loop, and inhibits SERCA through intra-membrane interactions. The cytoplasmic domain of PLN is in equilibrium between a helical, membrane-associated T state and an unfolded, membrane-dissociated R state. Here, I summarize the work to probe the structures of the T and R states and elucidate the role of the conformational equilibrium in regulation of SERCA. Using solution and solid state NMR in combination with biochemical assays I show that the structures of T and R state but not their relative populations are conserved in different lipid environments and sample conditions. Furthermore, the T/ R equilibrium has a central role in SERCA regulation and is crucial to relieve the inhibition of the enzyme. These findings provide new insights into SERCA/PLN function and offer a unique view of the role of conformational equilibria and multiple conformational states in membrane protein structure and function.
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    Development and testing of a protocol for computational prediction of 1H and 13C NMR chemical shifts and thermochemistry and reaction analysis of benzyne formation and trapping
    (2013-01) Marell, Daniel Joshua
    Elucidating structures of novel compounds and investigation of new reactions are two tasks that experimental organic chemists address on a frequent basis. The pursuit of these objectives can be rigorous and time-consuming. Of the methods employed in elucidating the structure of novel compounds, nuclear magnetic resonance (NMR) is by far the most widely applied. Investigation into new reactions may require any number of techniques to understand the reaction scope, kinetics, optimal conditions, mechanisms, etc. In both cases, the use of computational methods is well-suited to augment the experimentalist's data to guide and understand the system being investigated. A protocol for facilitating computational prediction of NMR chemical shifts was developed. Application to a set of natural products previously evaluated against computed NMR shifts, showed improved accuracy, through analysis of the corrected mean-absolute error (CMAE). The protocol was further employed successfully to aid in analysis of experimental spectra for compounds synthesized by collaborators where multiple diastereomers were possible. Graphing templates were also created to allow for rapid inspection of possible structures without more in-depth statistical analysis. Thermodynamic and mechanistic analysis on the formation and reaction of benzyne was also performed. Thermodynamic restrictions on the ring-size of fused benzynocycloalkanes were investigated. Additionally, analysis of the energetics and transition state geometries for small-molecule trapping (both intra and intermolecular) of benzyne are discussed.