Matching Items (3)

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The Characterization of Cannabidiol Amorphous Solid Dispersions

Description

Generating amorphous solid dispersions (ASDs) containing active pharmaceutical ingredients has become a favorable technique of emerging prominence to improve drug solubility and overall bioavailability. Cannabidiol (CBD) has now become a

Generating amorphous solid dispersions (ASDs) containing active pharmaceutical ingredients has become a favorable technique of emerging prominence to improve drug solubility and overall bioavailability. Cannabidiol (CBD) has now become a major focus in cannabinoid research due to its ability to serve as an anti-inflammatory agent, showing promising results in treating a wide array of debilitating diseases and pathologies. The following work provides evidence for generating homogenous glass phase amorphous solid dispersions containing 50% (w/w) up to 75% (w/w) CBD concentrations in the domain size of 2 – 5 nm. Concentrations up to 85% (w/w) CBD were concluded homogenous in the supercooled liquid phase in domain sizes of 20 – 30 nm. The results were obtained from polarized light microscopy (PLM), differential scanning calorimetry (DSC), as well as solution and solid-state NMR spectroscopy.

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Date Created
  • 2019

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Structural perspectives on glycosaminoglycan-binding proteins and their receptors

Description

Glycosaminoglycans (GAGs) are long chains of negatively charged sulfated polysaccharides. They are often found to be covalently attached to proteins and form proteoglycans in the extracellular matrix (ECM). Many proteins

Glycosaminoglycans (GAGs) are long chains of negatively charged sulfated polysaccharides. They are often found to be covalently attached to proteins and form proteoglycans in the extracellular matrix (ECM). Many proteins bind GAGs through electrostatic interactions. GAG-binding proteins (GBPs) are involved in diverse physiological activities ranging from bacterial infections to cell-cell/cell-ECM contacts. This thesis is devoted to understanding how interactions between GBPs and their receptors modulate biological phenomena. Bacteria express GBPs on surface that facilitate dissemination and colonization by attaching to host ECM. The first GBP investigated in this thesis is decorin binding protein (DBP) found on the surface of Borrelia burgdorferi, causative pathogens in Lyme disease. DBPs bind GAGs of decorin, a proteoglycan in ECM. Of the two isoforms, DBPB is less studied than DBPA. In current work, structure of DBPB from B. burgdorferi and its GAG interactions were investigated using solution NMR techniques. DBPB adopts a five-helical structure, similar to DBPA. Despite similar GAG affinities, DBPB has its primary GAG-binding site on the lysine-rich C terminus, which is different from DBPA. Besides GAGs, GBPs in ECM also interact with cell surface receptors, such as integrins. Integrins belong to a big family of heterodimeric transmembrane proteins that receive extracellular cues and transmit signals bidirectionally to regulate cell adhesion, migration, growth and survival. The second part of this thesis focuses on αM I-domain of the promiscuous integrin αMβ2 (Mac-1 or CD11b/CD18) and explores the structural mechanism of αM I-domain interactions with pleiotrophin (PTN) and platelet factor 4 (PF4), which are cationic proteins with high GAG affinities. After completing the backbone assignment of αM I-domain, paramagnetic relaxation enhancement (PRE) experiments were performed to show that both PTN and PF4 bind αM I-domain using metal ion dependent adhesion site (MIDAS) in an Mg2+ independent way, which differs from the classical Mg2+ dependent mechanism used by all known integrin ligands thus far. In addition, NMR relaxation dispersion analysis revealed unique inherent conformational dynamics in αM I-domain centered around MIDAS and the crucial C-terminal helix. These dynamic motions are potentially functionally relevant and may explain the ligand promiscuity of the receptor, but requires further studies.

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Date Created
  • 2019

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INVESTIGATING MECHANISMS OF TRANSIENT RECEPTOR POTENTIAL REGULATION WITH NUCLEAR MAGNETIC RESONANCE AND ROSETTA COMPUTATIONAL BIOLOGY

Description

The physiological phenomenon of sensing temperature is detected by transient

receptor (TRP) ion channels, which are pore forming proteins that reside in the

membrane bilayer. The cold and hot sensing TRP channels

The physiological phenomenon of sensing temperature is detected by transient

receptor (TRP) ion channels, which are pore forming proteins that reside in the

membrane bilayer. The cold and hot sensing TRP channels named TRPV1 and TRPM8

respectively, can be modulated by diverse stimuli and are finely tuned by proteins and

lipids. PIRT (phosphoinositide interacting regulator of TRP channels) is a small

membrane protein that modifies TRPV1 responses to heat and TRPM8 responses to cold.

In this dissertation, the first direct measurements between PIRT and TRPM8 are

quantified with nuclear magnetic resonance and microscale thermophoresis. Using

Rosetta computational biology, TRPM8 is modeled with a regulatory, and functionally

essential, lipid named PIP2. Furthermore, a PIRT ligand screen identified several novel

small molecular binders for PIRT as well a protein named calmodulin. The ligand

screening results implicate PIRT in diverse physiological functions. Additionally, sparse

NMR data and state of the art Rosetta protocols were used to experimentally guide PIRT

structure predictions. Finally, the mechanism of thermosensing from the evolutionarily

conserved sensing domain of TRPV1 was investigated using NMR. The body of work

presented herein advances the understanding of thermosensing and TRP channel function

with TRP channel regulatory implications for PIRT.

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Created

Date Created
  • 2018