Development Of Tools For Phosphosite-Specific Kinase Identification And Discovery Of Phosphatase Substrates,
2017
Wayne State University
Development Of Tools For Phosphosite-Specific Kinase Identification And Discovery Of Phosphatase Substrates, Pavithra Maheshani Dedigama Arachchige
Wayne State University Dissertations
Phosphorylation is a ubiquitous post translational modification implicated in many diseases, such as cancer. The phosphorylation status of cellular proteins is regulated by the activity of kinases and phosphatases. The biological significance of many phosphorylation events remain unknown because the methods to determine which kinase or phosphatase is responsible for phosphorylation are limited. Previously, we established kinase-catalyzed labeling where kinases accept γ-modified ATP analogs, such as ATP-arylazide and ATP-biotin, to label phosphoproteins. To study substrates of kinases and phosphatases, here we developed two new methods using kinase-catalyzed labeling. As one application, we developed K-CLASP (Kinase-catalyzed CrossLinking And Streptavdin Purification) to …
Design, Synthesis And Biological Evaluation Of Histone Deacetylase (Hdac) Inhibitors: Saha (Vorinostat) Analogs And Biaryl Indolyl Benzamide Inhibitors Display Isoform Selectivity,
2017
Wayne State University
Design, Synthesis And Biological Evaluation Of Histone Deacetylase (Hdac) Inhibitors: Saha (Vorinostat) Analogs And Biaryl Indolyl Benzamide Inhibitors Display Isoform Selectivity, Ahmed Negmeldin
Wayne State University Dissertations
HDAC proteins have emerged as interesting targets for anti-cancer drugs due to their involvement in cancers, as well as several other diseases. Several HDAC inhibitors have been approved by the FDA as anti-cancer drugs, including SAHA (suberoylanilide hydroxamic acid, Vorinostat). Unfortunately, SAHA inhibits most HDAC isoforms, which limit its use as a pharmacological tool and may lead to side effects in the clinic. In this work we were interested in developing isoform selective HDAC inhibitors, which may decrease or eliminate the side effects associated with non-selective inhibitors treatment. In addition, isoform selective HDAC inhibitors can be used as biological tools …
Development Of Chemical Tools To Investigate Protein S-Glutathionylation In Response To Metabolic Alteration,
2017
Wayne State University
Development Of Chemical Tools To Investigate Protein S-Glutathionylation In Response To Metabolic Alteration, Kusal Theekshana Gayan Samarasinghe
Wayne State University Dissertations
Oxidative stress is a common characteristic of age-related diseases such as vascular diseases, diabetes and cancer. Many diseases are known to be regulated by glutathionylation. Glutathionylation is referred to as the formation of disulfide bond between a protein cysteine and a glutathione. To understand the molecular mechanisms behind the disease initiation and progression, identification of such glutathionylated proteins is important. Even though existing methods have been widely used, several limitations of these methods hinder the identification of such proteins in disease conditions. Therefore, we developed a versatile chemical method that generates clickable glutathione inside the cells. In this method, we …
Ligand Binding Studies Of A Peptide Targeting Helix 69 Of 23s Rrna In Bacterial Ribosomes,
2017
Wayne State University
Ligand Binding Studies Of A Peptide Targeting Helix 69 Of 23s Rrna In Bacterial Ribosomes, Hyosuk Seo
Wayne State University Dissertations
In the development of finding a peptide targeting H69 of 23S rRNA in bacterial ribosomes, phage display was employed at pH 5.5, a buffer condition previously reported of H69 preferring a closed conformation. After sequencing, several peptides were chosen through sequence alignment, followed by preparation using solid-phase peptide synthesis. The peptides were characterized using MALDI-TOF and purified with HPLC. A truncated peptide TARHIY was selected from FID assay. Through binding studies using ESI-MS, SPR, BLItz, and NMR, the binding properties of the peptide to H69 were determined, such as binding affinity, stoichiometry, and interaction site. The peptide exhibited moderate binding …
The Development Of Chemical Methods To Discover Kinase Substrates And Map Cell Signaling With Gamma-Modified Atp Analog-Dependent Kinase-Catalyzed Phosphorylation,
2017
Wayne State University
The Development Of Chemical Methods To Discover Kinase Substrates And Map Cell Signaling With Gamma-Modified Atp Analog-Dependent Kinase-Catalyzed Phosphorylation, Dissanayaka Mudiyanselage Maheeka Madhubashini Embogama
Wayne State University Dissertations
Kinase-catalyzed phosphorylation plays an important role in cell physiology by regulating a myriad of cellular functions. Thus aberrant kinase activity is implicated in various diseases. Methods are needed to discover kinase substrates and map signaling pathways to explore biology and to help drug discovery. A few techniques are currently available to discover kinase substrate and map cell signaling. However, to augment kinase substrate discovery approaches, it is essential to develop alternative techniques. Pflum has recently discovered cosubstrate promiscuity of protein kinases with gamma-modified ATP analogs. Here, kinase-catalyzed biotinylation with ATP-biotin was used to develop novel tools to discover kinase substrates …
Ptpro Represses Erbb2-Driven Breast Oncogenesis By Dephosphorylation And Endosomal Internalization Of Erbb2.,
2017
George Washington University
Ptpro Represses Erbb2-Driven Breast Oncogenesis By Dephosphorylation And Endosomal Internalization Of Erbb2., H Dong, L Ma, J Gan, W Lin, Rakesh Kumar, +9 Additional Authors
Biochemistry and Molecular Medicine Faculty Publications
The plasma membrane-associated tyrosine phosphatase PTPRO is frequently transcriptionally repressed in cancers and signifies poor prognosis of breast cancer patients. In this study, deletion of Ptpro inMMTV-Erbb2 transgenic mice dramatically shortened the mammary tumor latency and accelerated tumor growth due to loss of Ptpro within the breast cancer cells but not in surrounding tissue as confirmed by hetero-transplantation studies. Both in vitro and in vivo data demonstrated that the phosphatase activity was required for the inactivation of ERBB2 and its downstream signaling. PTPRO regulated the phosphorylation status of ERBB2 at Y1248. Co-immunoprecipitation and proximity ligation assay (Duolink) indicated that …
Biomuta And Bioxpress: Mutation And Expression Knowledgebases For Cancer Biomarker Discovery,
2017
George Washington University
Biomuta And Bioxpress: Mutation And Expression Knowledgebases For Cancer Biomarker Discovery, Hayley Dingerdissen, John Torcivia-Rodriguez, Yu Hu, Ting-Chia Chang, Raja Mazumder, Robel Kashay
Biochemistry and Molecular Medicine Faculty Publications
Single-nucleotide variation and gene expression of disease samples represent important resources for biomarker discovery. Many databases have been built to host and make available such data to the community, but these databases are frequently limited in scope and/or content. BioMuta, a database of cancer-associated single-nucleotide variations, and BioXpress, a database of cancer-associated differentially expressed genes and microRNAs, differ from other disease-associated variation and expression databases primarily through the aggregation of data across many studies into a single source with a unified representation and annotation of functional attributes. Early versions of these resources were initiated by pilot funding for specific research …
Nanoparticle Targeting To The Central Nervous System,
2017
The University of Akron
Nanoparticle Targeting To The Central Nervous System, Emily Baker
Williams Honors College, Honors Research Projects
Drug delivery to the central nervous system is complicated by the blood-brain barrier, a vascular structure that prevents free diffusion of molecules into brain tissue. In this study, we examined the in vitro properties of a nanoparticle-based drug delivery system. We used several nanoparticle formulations, both LTP based and PLGA based, to test the cellular uptake and toxicity in microglial cells. Using immunofluorescence imaging, we show that LTP nanoparticles are taken up by microglia. We confirmed that our nanoparticle formulations are non-toxic by two cell viability assays. These results suggest that nanoparticle formulations may be a biocompatible method of delivering …
Dream Of A Conducive Learning Environment: One Dreem For All Medical Students!,
2017
Aga Khan University
Dream Of A Conducive Learning Environment: One Dreem For All Medical Students!, Rehana Rehman, Kulsoom Ghias, Syeda Sadia Fatima, Mehwish Hussain, Faiza Alam
Department of Biological & Biomedical Sciences
Objective: To compare students' response assessed by Dundee Ready Educational Environment Measure on the basis of the year of study, gender and pre-medical educational background.Methods: This cross-sectional study was carried out at the Aga Khan University Medical College, Karachi, Pakistan, from June 2014 to March 2015, and comprised medical students. The average scores of Dundee Ready Educational Environment Measure scales and subscales were compared between gender and educational background. Responses were also compared on the basis of year of study.Results: Of the 416 participants, 184 (44%) were males and 232 (56%) were females. There were 100 (24%) students in the …
A New Model For Phd Elective Course,
2017
Aga Khan University
A New Model For Phd Elective Course, H R. Ahmad, Satwat Hashmi, M H. Mahmood
Department of Biological & Biomedical Sciences
No abstract provided.
Investigating Medicinally Important Portein-Protein And Protein-Ligand Interactions : A Computational Approach,
2017
University of Richmond
Investigating Medicinally Important Portein-Protein And Protein-Ligand Interactions : A Computational Approach, Cooper Ashley Taylor
Honors Theses
Molecular dynamics (MD) simulations and computational chemistry allow for an atomistic understanding of protein-protein and protein-ligand binding motifs. Through the use of MD, medicinally relevant complexes can be examined in detail unattainable by experimental methods. Within this work, systems pertinent to both Alzheimer’s Disease and HIV-1 are probed and thoroughly sampled to help elucidate potential therapeutic pathways. We used molecular dynamics and free energy estimations to gauge the affinity for the binary and ternary complexes of KLC1, APP and JIP1, three proteins all believed to be involved in the propagation of Alzheimer’s Disease. Two areas of thought exist suggesting that …
Myeloperoxidase-Mediated Protein Lysine Oxidation Generates 2- Aminoadipic Acid And Lysine Nitrile In Vivo,
2017
Cleveland Clinic
Myeloperoxidase-Mediated Protein Lysine Oxidation Generates 2- Aminoadipic Acid And Lysine Nitrile In Vivo, Hongqiao Lin, Bruce S. Levison, Jennifer A. Buffa, Ying Huang, Xiaoming Fu, Zeneng Wang, Valentin Gogonea, Joseph A. Didonato, Stanley L. Hazen
Chemistry Faculty Publications
Recent studies reveal 2-aminoadipic acid (2-AAA) is both elevated in subjects at risk for diabetes and mechanistically linked to glucose homeostasis. Prior studies also suggest enrichment of protein-bound 2-AAA as an oxidative post-translational modification of lysyl residues in tissues associated with degenerative diseases of aging. While in vitro studies suggest redox active transition metals or myeloperoxidase (MPO) generated hypochlorous acid (HOCl) may produce protein-bound 2-AAA, the mechanism(s) responsible for generation of 2- AAA during inflammatory diseases are unknown. In initial studies we observed that traditional acid- or basecatalyzed protein hydrolysis methods previously employed to measure tissue 2-AAA can artificially generate …
Characterization Of Novel Borrelia Burgdorferi Transcripts Expressed During Tick And Mammalian Infection,
2017
University of Central Florida
Characterization Of Novel Borrelia Burgdorferi Transcripts Expressed During Tick And Mammalian Infection, Philip Adams
Electronic Theses and Dissertations
The purpose of this dissertation is to characterize the transcriptome of Borrelia (Borreliella) burgdorferi to discover novel transcripts, important for pathogenesis. As a spirochete and the etiological agent of Lyme disease, the foremost vector-borne bacterial infection in the world, B. burgdorferi fulfills a distinctive niche among bacterial pathogens. Persisting in the disparate environments of a tick vector and mammalian reservoirs, it is absolutely dependent on its hosts for transmission and nutrient acquisition. B. burgdorferi harbors a complex fragmented genome which is largely linear, unlike that of most prokaryotes, lacks an array of classically described metabolic genes, and contains an unusually …
Alpha-Synuclein: Insight Into The Hallmark Of Parkinson's Disease As A Target For Quantitative Molecular Diagnostics And Therapeutics,
2017
University of Central Florida
Alpha-Synuclein: Insight Into The Hallmark Of Parkinson's Disease As A Target For Quantitative Molecular Diagnostics And Therapeutics, Baggio A. Evangelista
Honors Undergraduate Theses
Parkinson’s disease (PD) is the second-most common neurodegenerative disease after Alzheimer’s disease. With 500,000 individuals currently living with Parkinson’s and nearly 60,000 new cases diagnosed each year, this disease causes significant financial burden on the healthcare system - amassing to annual expenditures totaling 200 billion dollars; predicted to increase through 2050. The disease phenotype is characterized by a combination of a resting tremor, bradykinesia, muscular rigidity, and depression due to dopaminergic neuronal death in the midbrain. The cause of the neurotoxicity has been largely discussed, with strong evidence suggesting that the protein, alpha-Synuclein, is a key factor. Under native conditions, …
Characterizing The Function Of The N-Terminal Domain Of Omi/Htra2,
2017
University of Central Florida
Characterizing The Function Of The N-Terminal Domain Of Omi/Htra2, Christine Nguyen
Honors Undergraduate Theses
The yeast two-hybrid system was used to isolate and characterize protein interactors of the N-terminal domain of the serine protease Omi/HtrA2 (high temperature requirement protein A2) encompassing amino acids 31-133. This large domain of Omi/HtrA2 is usually cleaved and removed through autoproteolysis to produce the mature form of the protein. Whether the N-terminal domain has any function after its removal is unknown. Omi/HtrA2 is involved in a variety of diseases including cancers, neurodegenerative disorders, and metabolic disorders, but thus far, it is assumed that its normal function is the degradation of specific substrates. To characterize any potential function of Omi/HtrA2’s …
A Family Of Genus-Specific Rnas In Tandem With Dna-Binding Proteins Control Expression Of The Bada Major Virulence Factor Gene In Bartonella Henselae,
2017
University of South Florida
A Family Of Genus-Specific Rnas In Tandem With Dna-Binding Proteins Control Expression Of The Bada Major Virulence Factor Gene In Bartonella Henselae, Nhan Tu, Ronan K. Carroll, Andy Weiss, Lindsey N. Shaw, Gael Nicolas, Sarah Thomas, Amorce Lima, Udoka Okaro, Burt Anderson
Molecular Biosciences Faculty Publications
Bartonella henselae is a gram-negative zoonotic bacterium that causes infections in humans including endocarditis and bacillary angiomatosis. B. henselae has been shown to grow as large aggregates and form biofilms in vitro. The aggregative growth and the angiogenic host response requires the trimeric autotransporter adhesin BadA. We examined the transcriptome of the Houston-1 strain of B. henselae using RNA-seq revealing nine novel, highly-expressed intergenic transcripts (Bartonella regulatory transcript, Brt1-9). The Brt family of RNAs is unique to the genus Bartonella and ranges from 194 to 203 nucleotides with high homology and stable predicted secondary structures. Immediately downstream of each …
Transposon-Mediated Stable Suppression Of Gene Expression In The Developing Chick Retina,
2017
Valparaiso University
Transposon-Mediated Stable Suppression Of Gene Expression In The Developing Chick Retina, Masaru Nakamoto, Chizu Nakamoto
Biology Faculty Publications
The embryonic chick has long been a favorite model system for in vivo studies of vertebrate development. However, a major technical limitation of the chick embryo has been the lack of efficient loss-of-function approaches for analyses of gene functions. Here, we describe a methodology in which a transgene encoding artificial microRNA sequences is introduced into embryonic chick retinal cells by in ovo electroporation and integrated into the genome using the Tol2 transposon system. We show that this methodology can induce potent and stable suppression of gene expression. This technique therefore provides a rapid and robust loss-of-function approach for studies of …
Exploring The Physics Of Proteins At Molecular Level By Neutron And X-Ray Scattering,
2017
Wayne State University
Exploring The Physics Of Proteins At Molecular Level By Neutron And X-Ray Scattering, Utsab Raj Shrestha
Wayne State University Dissertations
The protein structures revealed by the crystallographic studies have provided the valuable information over the years regarding their biological functions. However, such snapshots of protein fluctuations averaged over time may not be enough to fully capture the underlying biological phenomena. A deeper understanding of the protein dynamics is crucial for elucidating the structural pathways or the transition mechanism from the initial state to the final state necessary for regulating the physical and chemical processes. Hence, the biological activities and functions are mainly governed by the protein conformational dynamics. However, the direct correlation of a wide range of protein dynamics to …
Real-Time Investigation Of Bulky Lesion Bypass By Y-Family Dna Polymerase, Dpo4, Using Single Molecule Fret,
2017
Wayne State University
Real-Time Investigation Of Bulky Lesion Bypass By Y-Family Dna Polymerase, Dpo4, Using Single Molecule Fret, Pramodha Liyanage
Wayne State University Dissertations
DNA is constantly exposed to various DNA damaging agents that are generated by various internal and external sources. Some of this damage may not be able to be repaired by cellular machineries causing DNA replication to be blocked. Once the replication fork is blocked by a DNA adduct, damage tolerance DNA polymerases, mainly Y-family, are able to restore the DNA replication by synthesizing past the DNA adduct. Benzo[a]pyrene (B[a]P) is one of the most studied environmental carcinogens. It is known to make covalent DNA adducts after metabolic activation and the bulkiness of the B[a]P adducts impose a strong barrier to …
Frnk Regulatory Complex Formation With Fak Is Regulated By Erk Mediated Serine 217 Phosphorylation,
2017
Loyola University Chicago
Frnk Regulatory Complex Formation With Fak Is Regulated By Erk Mediated Serine 217 Phosphorylation, Taylor J. Zak
Dissertations
Focal adhesion kinase related non-kinase (FRNK) is an endogenous inhibitor of focal adhesion kinase (FAK) that has traditionally been used to inhibit FAK signaling in a variety of experiments and is also an important endogenous regulator of FAK signaling. More recently, FRNK has been shown to be of increasing importance in some pathologic conditions. Despite the increasing importance of FRNK, the molecular mechanism by which it functions remains unclear. In addition, FRNK contains several phosphorylation sites with unknown importance and function. Here I hypothesize that FRNK can inhibit FAK by binding directly to FAK within focal adhesions. Furthermore, I propose …
