Matching Items (5)
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- All Subjects: Protein
- Creators: Ros, Robert
Description
An animal's ability to produce protein-based silk materials has evolved independently in many different arthropod lineages, satisfying various ecological necessities. However, regardless of their wide range of uses and their potential industrial and biomedical applications, advanced knowledge on the molecular structure of silk biopolymers is largely limited to those produced by spiders (order Araneae) and silkworms (order Lepidoptera). This thesis provides an in-depth molecular-level characterization of silk fibers produced by two vastly different insects: the caddisfly larvae (order Trichoptera) and the webspinner (order Embioptera).
The molecular structure of caddisfly larval silk from the species Hesperophylax consimilis was characterized using solid-state nuclear magnetic resonance (ss-NMR) and Wide Angle X-ray Diffraction (WAXD) techniques. This insect, which typically dwells in freshwater riverbeds and streams, uses silk fibers as a strong and sticky nanoadhesive material to construct cocoons and cases out available debris. Conformation-sensitive 13C chemical shifts and 31P chemical shift anisotropy (CSA) information strongly support a unique protein motif in which phosphorylated serine- rich repeats (pSX)4 complex with di- and trivalent cations to form rigid nanocrystalline β-sheets. Additionally, it is illustrated through 31P NMR and WAXD data that these nanocrystalline structures can be reversibly formed, and depend entirely on the presence of the stabilizing cations.
Nanofiber silks produced by webspinners (order Embioptera) were also studied herein. This work addresses discrepancies in the literature regarding fiber diameters and tensile properties, revealing that the nanofibers are about 100 nm in diameter, and are stronger (around 500 MPa mean ultimate stress) than previous works suggested. Fourier-transform Infrared Spectroscopy (FT-IR), NMR and WAXD results find that approximately 70% of the highly repetitive glycine- and serine-rich protein core is composed of β-sheet nanocrystalline structures. In addition, FT-IR and Gas-chromatography mass spectroscopy (GC-MS) data revealed a hydrophobic surface coating rich in long-chain lipids. The effect of this surface coating was studied with contact angle techniques; it is shown that the silk sheets are extremely hydrophobic, yet due to the microstructural and nanostructural details of the silk surface, are surprisingly adhesive to water.
The molecular structure of caddisfly larval silk from the species Hesperophylax consimilis was characterized using solid-state nuclear magnetic resonance (ss-NMR) and Wide Angle X-ray Diffraction (WAXD) techniques. This insect, which typically dwells in freshwater riverbeds and streams, uses silk fibers as a strong and sticky nanoadhesive material to construct cocoons and cases out available debris. Conformation-sensitive 13C chemical shifts and 31P chemical shift anisotropy (CSA) information strongly support a unique protein motif in which phosphorylated serine- rich repeats (pSX)4 complex with di- and trivalent cations to form rigid nanocrystalline β-sheets. Additionally, it is illustrated through 31P NMR and WAXD data that these nanocrystalline structures can be reversibly formed, and depend entirely on the presence of the stabilizing cations.
Nanofiber silks produced by webspinners (order Embioptera) were also studied herein. This work addresses discrepancies in the literature regarding fiber diameters and tensile properties, revealing that the nanofibers are about 100 nm in diameter, and are stronger (around 500 MPa mean ultimate stress) than previous works suggested. Fourier-transform Infrared Spectroscopy (FT-IR), NMR and WAXD results find that approximately 70% of the highly repetitive glycine- and serine-rich protein core is composed of β-sheet nanocrystalline structures. In addition, FT-IR and Gas-chromatography mass spectroscopy (GC-MS) data revealed a hydrophobic surface coating rich in long-chain lipids. The effect of this surface coating was studied with contact angle techniques; it is shown that the silk sheets are extremely hydrophobic, yet due to the microstructural and nanostructural details of the silk surface, are surprisingly adhesive to water.
ContributorsAddison, John Bennett (Author) / Yarger, Jeffery L (Thesis advisor) / Holland, Gregory P (Thesis advisor) / Wang, Xu (Committee member) / Ros, Robert (Committee member) / Arizona State University (Publisher)
Created2014
Description
Proteins are a fundamental unit in biology. Although proteins have been extensively studied, there is still much to investigate. The mechanism by which proteins fold into their native state, how evolution shapes structural dynamics, and the dynamic mechanisms of many diseases are not well understood. In this thesis, protein folding is explored using a multi-scale modeling method including (i) geometric constraint based simulations that efficiently search for native like topologies and (ii) reservoir replica exchange molecular dynamics, which identify the low free energy structures and refines these structures toward the native conformation. A test set of eight proteins and three ancestral steroid receptor proteins are folded to 2.7Å all-atom RMSD from their experimental crystal structures. Protein evolution and disease associated mutations (DAMs) are most commonly studied by in silico multiple sequence alignment methods. Here, however, the structural dynamics are incorporated to give insight into the evolution of three ancestral proteins and the mechanism of several diseases in human ferritin protein. The differences in conformational dynamics of these evolutionary related, functionally diverged ancestral steroid receptor proteins are investigated by obtaining the most collective motion through essential dynamics. Strikingly, this analysis shows that evolutionary diverged proteins of the same family do not share the same dynamic subspace. Rather, those sharing the same function are simultaneously clustered together and distant from those functionally diverged homologs. This dynamics analysis also identifies 77% of mutations (functional and permissive) necessary to evolve new function. In silico methods for prediction of DAMs rely on differences in evolution rate due to purifying selection and therefore the accuracy of DAM prediction decreases at fast and slow evolvable sites. Here, we investigate structural dynamics through computing the contribution of each residue to the biologically relevant fluctuations and from this define a metric: the dynamic stability index (DSI). Using DSI we study the mechanism for three diseases observed in the human ferritin protein. The T30I and R40G DAMs show a loss of dynamic stability at the C-terminus helix and nearby regulatory loop, agreeing with experimental results implicating the same regulatory loop as a cause in cataracts syndrome.
ContributorsGlembo, Tyler J (Author) / Ozkan, Sefika B (Thesis advisor) / Thorpe, Michael F (Committee member) / Ros, Robert (Committee member) / Kumar, Sudhir (Committee member) / Shumway, John (Committee member) / Arizona State University (Publisher)
Created2011
Description
Proteins continually and naturally incur evolutionary selection through mutagenesis that optimizes their fitness, which is primarily determined by their function. It is known that allosteric regulation alters a protein's conformational dynamics leading to functional changes. We have computationally introduced a mutation at a predicted regulatory site of a short, 46 residue-long, protein interaction module composed of a WW domain and corresponding polyproline ligand (PDB id: 1k9r). The dynamic flexibility index (DFI) was computed for the binding site of the wild type and mutant WW domains to quantify the mutations effect on the rigidity of the binding pocket. DFI is used as a metric to quantify the resilience of a given position to perturbation along the chain. Using steered molecular dynamics (SMD), we also measure the effect of the point mutation on allosteric regulation by approximating the binding free energy of the system calculated using Jarzynski's Equality. Calculation of the DFI shows that the overall flexibility of the protein complex increases as a result of the distal point mutation. Total change in DFI percentile of the binding site showed a 0.067 increase suggesting an allosteric, loss of function mutation. Furthermore, we see that the change in the binding free energy is greater for that of the mutated complex supporting the idea that an increase in flexibility is correlated to a decrease in proteinlig and binding affinity. We show that sequence mutation of an allosteric site affects the mechanical stability and functionality of the binding pocket.
ContributorsMarianchuk, Tegan (Author) / Ozkan, Sefika (Thesis director) / Ros, Robert (Committee member) / Barrett, The Honors College (Contributor) / Department of Physics (Contributor)
Created2018-05
Description
Food production and consumption directly impacts the environment and human health. Protein in particular has significant cultural and health implications, and how people make decisions about what type of protein they eat has not been studied directly. Many decision tools exist to offer recommendations for seafood, but neglect livestock or plant protein. This study attempts to address these shortcomings in food decision science and tools by asking the questions: 1) What qualities of a dietary protein-based decision tool make it effective? 2) What do people consider when making decisions about what type of protein to consume? Using literature review, meta-analysis, and surveys, this study attempts to determine how the knowledge gained from answering these questions can be used to develop an electronic tool to engage consumers in making sustainable and healthy decisions about protein consumption. The data show that, given environmental and health information about the protein types, people in the sample of farmers market shoppers are more likely to purchase wild salmon and organically grown soybeans, and less likely to purchase grain-fed beef. However, the order of preference among the six types of protein did not change. Additional results suggest that there is a disconnect between consumers and sources of dietary protein, indicating a need for improved education. Inconsistency in labeling and information regarding protein types is a large source of confusion for consumers who participated in the survey, highlighting the need for transparency. Results of this study suggest that decisions tools may help improve decision making, but new ways of using them need to be considered to achieve this.
ContributorsGeren, Sarah (Author) / Gerber, Leah (Thesis advisor) / Minteer, Ben (Committee member) / Wentz, Elizabeth (Committee member) / Arvai, Joseph (Committee member) / Arizona State University (Publisher)
Created2015
Description
My research centers on the design and fabrication of biomolecule-sensing devices that combine top-down and bottom-up fabrication processes and leverage the unique advantages of each approach. This allows for the scalable creation of devices with critical dimensions and surface properties that are tailored to target molecules at the nanoscale.
My first project focuses on a new strategy for preparing solid-state nanopore sensors for DNA sequencing. Challenges for existing nanopore approaches include specificity of detection, controllability of translocation, and scalability of fabrication. In a new solid-state pore architecture, top-down fabrication of an initial electrode gap embedded in a sealed nanochannel is followed by feedback-controlled electrochemical deposition of metal to shrink the gap and define the nanopore size. The resulting structure allows for the use of an electric field to control the motion of DNA through the pore and the direct detection of a tunnel current through a DNA molecule.
My second project focuses on top-down fabrication strategies for a fixed nanogap device to explore the electronic conductance of proteins. Here, a metal-insulator-metal junction can be fabricated with top-down fabrication techniques, and the subsequent electrode surfaces can be chemically modified with molecules that bind strongly to a target protein. When proteins bind to molecules on either side of the dielectric gap, a molecular junction is formed with observed conductances on the order of nanosiemens. These devices can be used in applications such as DNA sequencing or to gain insight into fundamental questions such as the mechanism of electron transport in proteins.
My first project focuses on a new strategy for preparing solid-state nanopore sensors for DNA sequencing. Challenges for existing nanopore approaches include specificity of detection, controllability of translocation, and scalability of fabrication. In a new solid-state pore architecture, top-down fabrication of an initial electrode gap embedded in a sealed nanochannel is followed by feedback-controlled electrochemical deposition of metal to shrink the gap and define the nanopore size. The resulting structure allows for the use of an electric field to control the motion of DNA through the pore and the direct detection of a tunnel current through a DNA molecule.
My second project focuses on top-down fabrication strategies for a fixed nanogap device to explore the electronic conductance of proteins. Here, a metal-insulator-metal junction can be fabricated with top-down fabrication techniques, and the subsequent electrode surfaces can be chemically modified with molecules that bind strongly to a target protein. When proteins bind to molecules on either side of the dielectric gap, a molecular junction is formed with observed conductances on the order of nanosiemens. These devices can be used in applications such as DNA sequencing or to gain insight into fundamental questions such as the mechanism of electron transport in proteins.
ContributorsSadar, Joshua Stephen (Author) / Qing, Quan (Thesis advisor) / Lindsay, Stuart (Committee member) / Vaiana, Sara (Committee member) / Ros, Robert (Committee member) / Arizona State University (Publisher)
Created2019