Life Sciences Commons^™

Structural And Functional Studies Of The Papain-Like Protease 2 From Mouse Hepatitis Virus, Yafang Chen

Open Access Dissertations

Our goal is to establish a system to investigate how the deubiquitinating (DUB) and deISGylating activities of coronavirus (CoV) papain-like protease domains (PLPs) are involved in virus immune evasion. To this end, we chose PLP2 from mouse hepatitis virus (MHV) as our target of study because MHV has historically served as a model system for the study of CoVs, and it has undeniable advantage of ease in culturing in comparison to human coronaviruses.

It is reported here the expression and purification of a region of MHV nsp3 that contains the catalytic core of the PLP2 domain and its neighboring domains. …

Elucidating The Role Of Hausp Ubiquitin Like Domains In The Catalytic Function Of Usp7, Anuj Patel, Nicole Davis, Andrew Mesecar

The Summer Undergraduate Research Fellowship (SURF) Symposium

Ubiquitin specific proteases (USPs) are a class of enzymes involved in myriad cellular processes. One USP of great interest due to its oncogenic properties is USP7. In normal conditions USP7 is closely regulated due to its responsibility for destabilizing the tumor suppressor, p53, through the deubiquitination of MDM2. In multiple myeloma cases, it appears the regulation of USP7 subsides, as it is largely overexpressed, leading to the inappropriate degradation of p53. Inhibition of USP7 could, therefore, prove a viable target for cancer therapy. A greater understanding of USP7’s function and structure can lead to more insight into how this enzyme …

The Effect Of Macromolecular Crowding On The Structure Of The Protein Complex Superoxide Dismutase, Ajith Rathnaweera Rajapaksha Mudalige

Open Access Dissertations

Biological environments contain between 7 - 40% macromolecules by volume. This reduces the available volume for macromolecules and elevates the osmotic pressure relative to pure water. Consequently, biological macromolecules in their native environments tend to adopt more compact and dehydrated conformations than those in vitro. This effect is referred to as macromolecular crowding and constitutes an important physical difference between native biological environments and the simple solutions in which biomolecules are usually studied.^ We used small angle scattering (SAS) to measure the effects of macromolecular crowding on the size of a protein complex, superoxide dismutase (SOD). Crowding was induced using …

A Novel In Vivo Tumor Oxygen Profiling Assay: Combining Functional And Molecular Imaging With Multivariate Mathematical Modeling, Chung-Wein Lee

Open Access Dissertations

Purpose: The objective of this study is to develop and test a novel high spatio-temporal in vivo assay to quantify tumor oxygenation and hypoxia. The assay implements a biophysical model of oxygen transport to fuse parameters acquired from in vivo functional and molecular imaging modalities. ^ Introduction: Tumor hypoxia plays an important role in carcinogenesis. It triggers pathological angiogenesis to supply more oxygen to the tumor cells and promotes cancer cell metastasis. Preclinical and clinical evidence show that anti-angiogenic treatment is capable of normalizing the tumor vasculature both structurally and functionally. The resulting normalized vasculature provides a more efficient and …

Quantitative Mrna Detection With Advanced Nonlinear Microscopy, Jing Liu

Open Access Dissertations

Cell-specific information on quantity and localization of key mRNA transcripts in single-cell level are critical to the assessment of cancer risk, therapy efficacy, and effective prevention strategies. While current techniques are not capable to visualize single mRNA transcript beyond the diffraction limit. In this thesis, two nonlinear technologies, second harmonic super-resolution microscopy (SHaSM) and transient absorption microscopy (TAM), are developed to detect and quantify single Human edimer receptor 2 (Her2) mRNA transcripts. The SHaSM is used to detect single mRNA transcript beyond the diffraction limit, while the TAM is employed to detect mRNA without the interference of fluorescence background. The …

Magnetic Manipulation And Multimodal Imaging For Single Cell Direct Mechanosensing, Robert L. Wilson

Open Access Theses

The study of internal mechanics of single cells is paramount to understand mechanisms of mechanoregulation. External loading and cell-mediated force generation result in changes in cell shape, rheology, and the deformation of subcellular structures such as the nucleus. Moreover, alterations in the processes that regulate these responses have been further correlated to specific pathologies. Cellular deformation is often studied through application of forces in the environment of the cell, relying on strain and stress transfer through focal adhesions and the cytoskeletal system. However, the transfer of these external forces to internal mechanics can introduce uncertainties in the interpretation of subcellular …

Biochemical Investigation Of The Ubiquitin Carboxyl-Terminal Hydrolase Family, Joseph Rashon Chaney

Open Access Dissertations

The proteasome is the machinery in eukaryotic cells that degrades protein and recycles the amino acids. Protein degradation is a highly regulated process which starts by the attachment of chains of ubiquitin, which serves as a tag that marks a protein for degradation. This function involves the work of several proteins at the proteasome that work either as ubiquitin chaperones, ubiquitin binders or cleave ubiquitin from the protein that is to be degraded. As this is a highly regulated process, various irregularities can have deleterious effects including the onset of disease, including cardiovascular, cancer, and neurological. ^ The focus of …

Development Of Experimental And Instrumental Systems To Study Biological Systems, Amanda J Hemphill

Open Access Dissertations

Chapters 1-4 of this thesis describes the development of an experimental system to measure diffusion-limited reaction kinetics in a biological environment. About 100 years ago, the relationship between reaction rate and diffusion in homogenous solution, ie water or buffer, was described as a linear relationship by Smoluchowski. Applying this theory naively would suggest that since the diffusion coefficients drop by factors of 4-100 then the rates of reaction would drop by the same amount. However, recent theory and simulations suggest that this does not hold. Even though biological diffusion coefficients drop to 0.1-20% of that in buffer, these recent studies …

Structural And Biophysical Analysis Of The Proteasomal Deubiquitinase, Uch37, Marie Elizabeth Morrow

Open Access Dissertations

Ubiquitin carboxyl-terminal hydrolase 37, or UCH37, is a deubiquitinating enzyme associated with the 26S proteasome, the primary protein degradation machinery in eukaryotic cells. UCH37 is responsible for the disassembly of polymeric ubiquitin chains, or polyubiquitin, which have been ligated onto proteins in order to target them for degradation. The 26S utilizes two associated deubiquitinating enzymes, UCH37 and USP14, and one intrinsic, Rpn11, to remove polyubiquitin chains from substrate proteins as they are unfolded and translocated into the proteolytic core of the proteasome, where proteins are cleaved into small peptides and then released for recycling by the cell. UCH37 associates with …

Evaluating Breast Cancer Stem Cell Response To Antiangiogenic Therapy, Connor J. Holloway

Open Access Theses

Angiogenic inhibitors function by blocking tumor cell signals used to recruit host tissue vasculature to the tumor site, thereby depriving the cancer of the nutriment needed for further expansion. The development and implementation of angiogenic inhibitors in conjunction with standard chemotherapy agents has increased progression-free survival but not overall patient survival. It is hypothesized that chronic exposure to large doses of AAT drugs worsens hypoxic conditions within the tumor mass, selectively stimulating aggressive cancer stem cell populations to grow and proliferate. ^ In this study, the expression of the CSC biomarkers ALDH1, DLL1, and EpCAM were evaluated in breast cancer …