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Full-Text Articles in Life Sciences

The Effects Of Acidosis On Calcium Dependent Binding Of A Single Crossbridge, Matthew Unger Oct 2019

The Effects Of Acidosis On Calcium Dependent Binding Of A Single Crossbridge, Matthew Unger

Masters Theses

Intracellular acidosis is a putative agent of skeletal muscle fatigue, in part, because acidosis depresses the calcium (Ca2+) sensitivity and force production of muscle (18, 50). However, the molecular mechanisms behind this depression in Ca2+ sensitivity and force production are unknown. This gap in knowledge poses a significant challenge in generating a complete understanding of the fatigue process. To close this gap, the ability of myosin to bind to a single actin filament was measured under acidic conditions, in a laser trap assay, with and without regulatory proteins. Decreasing pH from 7.4 to 6.5 reduced the ...


Multiscale Simulations Of Intrinsically Disordered Proteins, Xiaorong Liu Jul 2019

Multiscale Simulations Of Intrinsically Disordered Proteins, Xiaorong Liu

Doctoral Dissertations

Intrinsically disordered proteins (IDPs) lack stable secondary and/or tertiary structures under physiological conditions. The have now been recognized to play important roles in numerous biological processes, particularly cellular signaling and regulation. Mutation of IDPs are frequently associated with human diseases, such as cancers and neuron degenerative diseases. Therefore, it is important to understand the structure, dynamics, and interactions of IDPs, so as to establish the mechanistic basis of how intrinsic disorder mediates versatile functions and how such mechanisms may fail in human diseases. However, the heterogeneous structural ensembles of IDPs are not amenable to high resolution characterization solely through ...


Observing The Molecular Basis Of Thin Filament Activation With A Three Bead Laser Trap Assay, Thomas Longyear Mar 2018

Observing The Molecular Basis Of Thin Filament Activation With A Three Bead Laser Trap Assay, Thomas Longyear

Doctoral Dissertations

Muscle contracts after calcium (Ca++) is released into the muscle cell, resulting from a cascade of events which result in myosin, the molecular motor of muscle, to produce force and motion. Myosin cyclically binds to a regulated thin filament, using the chemical energy of ATP to produce force and motion. Perturbations in muscle, such as a build-up of metabolic by-products or point mutations in key contractile proteins, can inhibit these functions in both skeletal and cardiac muscle either acutely or chronically. Despite the many years we have studied skeletal and cardiac muscle, we still do not have a clear picture ...


The Role Of The Metallochaperone Hypa In The Acid Survival And Activities Of Nickel Enzymes In Helicobacter Pylori, Heidi Hu Mar 2018

The Role Of The Metallochaperone Hypa In The Acid Survival And Activities Of Nickel Enzymes In Helicobacter Pylori, Heidi Hu

Doctoral Dissertations

Helicobacter pylori is a bacterium that has colonized the human gastric mucosa of over 50% of the world population. Persistent infection can cause gastritis, peptic ulcers, and cancers. The ability of H. pylori to colonize the acidic environment of the human stomach is dependent on the activity of the nickel containing enzymes, urease and NiFe-hydrogenase. The nickel metallochaperone, HypA, was previously shown to be required for the full activity of both enzymes. In addition to a Ni-binding site, HypA also contains a structural Zn site, which has been characterized to alter its averaged structure depending on pH and the presence ...


Dissecting Molecular Pathways That Ensure Proper Chromosome Segregation And Cell Division, Anna Ye Nov 2017

Dissecting Molecular Pathways That Ensure Proper Chromosome Segregation And Cell Division, Anna Ye

Doctoral Dissertations

Equal segregation of the genome is a prerequisite for cell survival. During cell division the duplicated DNA is compacted into chromosomes and a multi-protein macrostructure, known as the kinetochore (Kt), is assembled on each copy of compacted DNA. Simultaneously, the mitotic spindle, which is made up of microtubules (MTs), is built to facilitate the equal distribution of the chromosomes between the resulting daughter cells. Kinetochores mediate the interaction between the MTs and the chromosomes, properly positioning them for segregation. To ensure that the DNA is equally divided in every cell division, cells have built a surveillance system to detect any ...


Pore Forming Protein Assembly And The Use In Nanopore Sensing: A Study On E. Coli Proteins Clya And Ompg, Monifa Fahie Nov 2017

Pore Forming Protein Assembly And The Use In Nanopore Sensing: A Study On E. Coli Proteins Clya And Ompg, Monifa Fahie

Doctoral Dissertations

Pore forming proteins are typically the proteins that form channels in membranes. They have several roles ranging from molecule transport to triggering the death of a cell. This work focuses on two E. coli pore forming proteins that have vastly differing roles in nature. Outer membrane protein G (OmpG) is an innocuous β-barrel porin while Cytolysin A (ClyA) is an α-helical pore forming toxin. For OmpG we probed its potential to be a nanopore sensor for protein detection and quantification. A small high affinity ligand, biotin, was covalently attached to loop 6 of OmpG and used to capture biotin-binding proteins ...


Regulation Of Katanin Activity On Microtubules, Madison A. Tyler Oct 2017

Regulation Of Katanin Activity On Microtubules, Madison A. Tyler

Masters Theses

The cytoskeleton is a dynamic network of microtubules constantly being reorganized to meet the spatiotemporal demands of the cell. Microtubules are organized into subcellular highways to control cell processes such as cell division, cargo transport, and neuronal development and maintenance. Reorganization of this intricate network is tightly regulated by various stabilizing and destabilizing microtubule-associated proteins that decorate the network. Katanin p60 is a microtubule destabilizing enzyme from the ATPases Associated with various Activities (AAA+) family. It can both sever and depolymerize microtubules. In order to sever microtubules, katanin recognizes the tubulin carboxy-terminal tails (CTTs) and hydrolyzes ATP. Using super-resolution microscopy ...


Vitreous Gel Physics, Svetlana Morozova Jul 2017

Vitreous Gel Physics, Svetlana Morozova

Doctoral Dissertations

The transparent vitreous, which fills the posterior cavity of the eye, is incredibly engineered. The charged polyelectrolyte hyaluronic acid (HA) network swells to maintain the pressure in the eye, while stiff collagen type II bundles help absorb any external mechanical shock. Our investigations have contributed to a few key developments related to the physical properties of the vitreous:

(1) The stiff collagen network that supports the soft gel network is self-assembled from single triple-helix collagen proteins. Electrostatic interactions drive this assembly, such that the size and concentration are optimized at physiological salt concentrations. The width of the assemblies remarkably changes ...


X-Ray Characterization Of Mesophases And Phase Transitions Of Dna Analogues In Solutions, Mustafa Selcuk Yasar Nov 2016

X-Ray Characterization Of Mesophases And Phase Transitions Of Dna Analogues In Solutions, Mustafa Selcuk Yasar

Doctoral Dissertations

We think of DNA as double-stranded helices (duplex), but the polymer exists in many conformations. Several triplex and quadruplex DNA structures can be formed in laboratory settings and exist in nature. This thesis first provides a brief description of the nature of the order in arrays of duplex DNA under biologically relevant molecular crowding conditions. Then we compare the duplex DNA mesophases with the corresponding liquid crystalline phase behavior of the triplex and quadruplex DNA analogues. In particular, we focus on G-quadruplexes.

Observed in the folds of guanine-rich oligonucleotides, G-quadruplex structures are based on G-quartets formed by hydrogen bonding and ...


Investigating The Impact Of Small Molecule Ligands And The Proteostasis Network On Protein Folding Inside The Cell, Karan Hingorani Nov 2016

Investigating The Impact Of Small Molecule Ligands And The Proteostasis Network On Protein Folding Inside The Cell, Karan Hingorani

Doctoral Dissertations

The folded forms of most proteins are critical to their functions. Despite the complexity of the cellular milieu and the presence of high-risk deleterious interactions, there is a high level of fidelity observed in the folding process for entire proteomes. Two important reasons for this are the presence of the quality control machinery consisting of chaperones and degradation enzymes that work jointly to optimize the population of the folded state and interaction partners that re-enforce the functional state and add to the competitive advantage of an organism. While substantial effort has been directed to understand protein folding and interactions in ...


Characterizing The Inhibition Of Katanin Using Tubulin Carboxy-Terminal Tail Constructs, Corey E. Reed Nov 2016

Characterizing The Inhibition Of Katanin Using Tubulin Carboxy-Terminal Tail Constructs, Corey E. Reed

Masters Theses

Understanding how the cellular cytoskeleton is maintained and regulated is important to elucidate the functions of many structures such as the mitotic spindle, cilia and flagella. Katanin p60, microtubule-severing enzymes from the ATPase associated with cellular activities (AAA+) family, has previously been shown in our lab to be inhibited by free tubulin as well as α- and β-tubulin carboxy-terminal tail (CTT) constructs. Here we investigate the inhibition ability of several different tubulin CTT sequences. We quantify the effect of the addition of these constructs on the severing and binding activity of katanin. We find that some constructs inhibit katanin better ...


Dynamics Of Microtubule Networks With Antiparallel Crosslinkers, Kasimira T. Stanhope Jul 2016

Dynamics Of Microtubule Networks With Antiparallel Crosslinkers, Kasimira T. Stanhope

Masters Theses

Microtubules are the most rigid element of the cytoskeleton. They are responsible for the structure of cells and make up the tracks for intracellular cargo transport. Interactions between microtubules, motor proteins, and microtubule-associated proteins drive important mechanisms in the cell, such as cell division, cell motility, cell homeostasis, and cell signaling. I seek to understand how such complex, energy-consuming non-equilibrium biological networks self-organize by studying in vitro microtubules bundled by microtubule-associated protein 65 (MAP65), in kinesin-1 gliding assays. I found that large networks can break into smaller, cell-like networks that can mimic types of cell motility. Dynamics of these networks ...


Biophysical Characterization Of Katanin’S Regulation Of Microtubules, Megan E. Bailey Nov 2015

Biophysical Characterization Of Katanin’S Regulation Of Microtubules, Megan E. Bailey

Doctoral Dissertations

Microtubules, as an essential part of the cytoskeleton, require proper function as well as correct spatial and temporal localization. In order to achieve correct organization, microtubule-associated proteins (MAPs) regulate microtubule dynamics. Katanin, a known microtubule-severing enzyme from the AAA family of proteins, plays a role in regulating microtubules, but the mechanisms of microtubule control and the mechanism of severing activity remain to be elucidated.

In the following studies I examine mechanisms of katanin-based regulation of microtubule dynamics using a single molecule biophysics approach. I use this simplified in vitro approach to change specific parameters to investigate how katanin targets microtubules ...


Protein Behavior Directed By Heparin Charge And Chain Length, Burcu Baykal Minsky Aug 2014

Protein Behavior Directed By Heparin Charge And Chain Length, Burcu Baykal Minsky

Doctoral Dissertations

Glycosaminoglycans (GAGs), highly charged biological polyelectrolytes, are of growing importance as biomaterials and pharmaceutical drugs due to their immense range of physiological functions. They bind to many proteins; however, the degree of structural selectivity in GAG-protein interactions is largely unknown .Our studies have focused on the importance of heparin (a model GAG) charge and chain length in protein binding in order to explore its potential applications in biofunctional tissue scaffold materials, as polysaccharide drugs in anticoagulation, and as inhibitory agents in protein aggregation. We used electrospray ionization mass spectrometry, capillary electrophoresis, size exclusion chromatography, dynamic/static light scattering and electrostatic ...


Structural Studies Of Membrane-Assembled Popd And Popb, The Pseudomonas Aeruginosa Type 3 Secretion Translocators, Fabian B Romano Chernac Sep 2012

Structural Studies Of Membrane-Assembled Popd And Popb, The Pseudomonas Aeruginosa Type 3 Secretion Translocators, Fabian B Romano Chernac

Doctoral Dissertations 1896 - February 2014

Transport of proteins across membranes is essential during many stages of pathogen infection and colonization of human cells. Many Gram-negative pathogens use a Type 3 Secretion (T3S) system to inject proteins into the target cell during infection. Substantial genetic and biochemical evidence suggest that proteins are translocated across the host plasma membrane through a proteinaceous pore or translocon formed by two bacterial secreted proteins: the T3S translocators. Despite its key role in pathogenesis, virtually nothing is known about the assembly mechanism, structure, and composition of this critical transmembrane complex.

To this end, a cell-free system for the structural and functional ...