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Articles 1 - 30 of 57
Full-Text Articles in Life Sciences
Nucleobase-Modified Nucleosides And Nucleotides: Applications In Biochemistry, Synthetic Biology, And Drug Discovery, Anthony J. Berdis
Nucleobase-Modified Nucleosides And Nucleotides: Applications In Biochemistry, Synthetic Biology, And Drug Discovery, Anthony J. Berdis
Chemistry Faculty Publications
DNA is often referred to as the "molecule of life " since it contains the genetic blueprint for all forms of life on this planet. The core building blocks composing DNA are deoxynucleotides. While the deoxyribose sugar and phosphate group are ubiquitous, it is the composition and spatial arrangement of the four natural nucleobases, adenine (A), cytosine (C), guanine (G), and thymine (T), that provide diversity in the coding information present in DNA. The ability of DNA to function as the genetic blueprint has historically been attributed to the formation of proper hydrogen bonding interactions made between complementary nucleobases. However, …
Circadian Clock Controls Rhythms In Ketogenesis By Interfering With Ppar Alpha Transcriptional Network, Volha Mezhnina, Oghogho P. Ebeigbe, Nikkhil Velingkaar, Allan Poe, Yana I. Sandlers, Roman Kondratov
Circadian Clock Controls Rhythms In Ketogenesis By Interfering With Ppar Alpha Transcriptional Network, Volha Mezhnina, Oghogho P. Ebeigbe, Nikkhil Velingkaar, Allan Poe, Yana I. Sandlers, Roman Kondratov
Biological, Geological, and Environmental Faculty Publications
Ketone bodies are energy-rich metabolites and signaling molecules whose production is mainly regulated by diet. Caloric restriction (CR) is a dietary intervention that improves metabolism and extends longevity across the taxa. We found that CR induced high -amplitude daily rhythms in blood ketone bodies (beta-hydroxybutyrate [beta OHB]) that correlated with liver beta OHB level. Time-restricted feeding, another periodic fasting-based diet, also led to rhythmic beta OHB but with reduced amplitude. CR induced strong circadian rhythms in the expression of fatty acid oxidation and ketogenesis genes in the liver. The transcriptional factor peroxisome-proliferator-activated-receptor alpha (PPAR alpha) and its transcriptional target hepatokine …
Targeting Heat Shock 27 Kda Protein Induces Androgen Receptor Degradation, Yaxin Li
Targeting Heat Shock 27 Kda Protein Induces Androgen Receptor Degradation, Yaxin Li
ETD Archive
Glioblastoma (GBM) is the most common and aggressive brain tumor, with very poor prognosis. Androgen receptor (AR) plays a significant role in the progression of GBM, and anti-androgen agents have the potential to be used for the treatment of GBM. However, AR mutation commonly happens in GBM, which makes the anti-androgen agents less effective. Heat shock 27 kDa protein (HSP27) is a well-documented chaperone protein to stabilize AR. Inhibition of HSP27 results in AR degradation regardless the mutation status of AR, which makes HSP27 a good target to abolish AR in GBM. Identified compound I ((N-(3-((2,5-dimethoxybenzyl)oxy)-4-(methylsulfonamido) phenyl)-4-methoxybenzamide) inhibits GBM cell …
Mechanisms Of Telomere Maintenance In Trypanosoma Brucei, M A G G. Rabbani
Mechanisms Of Telomere Maintenance In Trypanosoma Brucei, M A G G. Rabbani
ETD Archive
Telomeres are a nucleoprotein structure at the end of the chromosome and are essential for genome integrity and chromosome stability. Telomere lengths are primarily maintained by a telomerase-mediated pathway but can be maintained by a homologous recombination-mediated pathway. However, detailed mechanisms of telomere maintenance are still unclear in many eukaryotes, including an important human pathogen, Trypanosoma brucei. Telomeres can be elongated by telomerase in T. brucei, a causative agent of fatal sleeping sickness in humans and nagana in cattle. T. brucei evades host immune response by regularly switching its major surface antigen, variant surface glycoprotein (VSG), a process known as …
Analysis Of Oxygen-18 Labeled Phosphate To Study Positional Isotope Experiments Using Lc-Qtof-Ms, Sujatha Chilakala, Iteen Cheng, Ireen Lee, Yan Xu
Analysis Of Oxygen-18 Labeled Phosphate To Study Positional Isotope Experiments Using Lc-Qtof-Ms, Sujatha Chilakala, Iteen Cheng, Ireen Lee, Yan Xu
Chemistry Faculty Publications
A method is proposed in this paper for the determination of oxygen-18 labeled phosphate so that positional isotope experiments using sensitive and rapid liquid chromatography–QTOF–mass spectrometry (LC-QTOF-MS) experiments can be carried out. The positional isotope exchange technique is a useful tool in understanding the mechanisms and kinetics of many enzyme-catalyzed reactions. Detection of the positions and concentration of these exchanged isotopes is the key. Gas chromatography–mass spectrometry (GC-MS) and nuclear magnetic resonance imaging are commonly used analytical techniques for measurement of 18O/16O, 31P and 15N isotope enrichment. Since these techniques either require a time-consuming derivatization …
Amino Acids Profiling For The Diagnosis Of Metabolic Disorders, Yana Sandlers
Amino Acids Profiling For The Diagnosis Of Metabolic Disorders, Yana Sandlers
Chemistry Faculty Publications
Inborn errors of metabolism (IEM) represent a group of inherited diseases in which genetic defect leads to the block on a metabolic pathway, resulting in a single enzyme dysfunction. As a downstream consequence of the residual or full loss of the enzymatic activity, there is an accumulation of toxic metabolites in the proximity of the metabolic block and/or a deficiency of an essential metabolic product which leads to the clinical presentation of the disease. While individually IEMs are rare, a collectively estimated incidence of metabolic inherited disorders is 1:800. The genetic basis of IEMs can involve abnormalities such as point …
Manganese Oxide/Hemin-Functionalized Graphene As A Platform For Peroxynitrite Sensing, Haitham F. Kalil, Shaimaa Maher, Tiyash Bose, Mekki Bayachou
Manganese Oxide/Hemin-Functionalized Graphene As A Platform For Peroxynitrite Sensing, Haitham F. Kalil, Shaimaa Maher, Tiyash Bose, Mekki Bayachou
Chemistry Faculty Publications
Peroxynitrite (ONOO−, PON) is a powerful oxidizing agent generated in vivo by the diffusion-limited reaction of nitric oxide (NO) and superoxide (O2˙−) radicals. Under oxidative stress, cumulated peroxynitrite levels are associated with chronic inflammatory disorders and other pathophysiological conditions. The accurate detection of peroxynitrite in biological systems is important, not only to understand the genesis and development of diseases, but also to explore and design potential therapeutics. Herein, a manganese oxide/hemin-modified graphene interface is explored as a platform for peroxynitrite amperometric detection. Hemin-functionalized reduced graphene oxide was further modified with manganese oxide nanoparticles to provide a …
N-Glycosylation In The Protease Domain Of Trypsin-Like Serine Proteases Mediates Calnexin-Assisted Protein Folding, Hao Wang, Shuo Li, Juejin Wang, Shenghan Chen, Xue-Long Sun, Qingyu Wu
N-Glycosylation In The Protease Domain Of Trypsin-Like Serine Proteases Mediates Calnexin-Assisted Protein Folding, Hao Wang, Shuo Li, Juejin Wang, Shenghan Chen, Xue-Long Sun, Qingyu Wu
Chemistry Faculty Publications
Trypsin-like serine proteases are essential in physiological processes. Studies have shown that N-glycans are important for serine protease expression and secretion, but the underlying mechanisms are poorly understood. Here, we report a common mechanism of N-glycosylation in the protease domains of corin, enteropeptidase and prothrombin in calnexin-mediated glycoprotein folding and extracellular expression. This mechanism, which is independent of calreticulin and operates in a domain-autonomous manner, involves two steps: direct calnexin binding to target proteins and subsequent calnexin binding to monoglucosylated N-glycans. Elimination of N-glycosylation sites in the protease domains of corin, enteropeptidase and prothrombin inhibits corin and enteropeptidase cell surface …
Evaluation Of A Small Molecule Agonist Of Epha2 Receptor Tyrosine Kinase And Copalic Acid Analogs As Prostate Cancer Therapeutics, Nethrie Idippily
Evaluation Of A Small Molecule Agonist Of Epha2 Receptor Tyrosine Kinase And Copalic Acid Analogs As Prostate Cancer Therapeutics, Nethrie Idippily
ETD Archive
Project I: Chemotherapeutic drugs have many side effects that are undesirable and are highly toxic. Therefore, there is a growing need for the development of drugs with enhanced efficacy, specificity, and potency to provide cancer patients with a better prognosis. It was discovered that a member of the Receptor Tyrosine Kinase family, EphA2, may prove to be a viable target in developing anti-cancer agents. In the presence of its ligand, EphA2 receptor is responsible for apoptotic and anti-migratory activity. However, in the absence of ligand, EphA2 is able to stimulate cell migration and therefore tumorigenic activity. These conflicting roles of …
Quantitative Analysis Of Bleomycin In Rat Plasma By Lc-Ms/Ms, Huawen Li
Quantitative Analysis Of Bleomycin In Rat Plasma By Lc-Ms/Ms, Huawen Li
ETD Archive
Bleomycin is the most commonly used compound in its group of antineoplastic drugs. It works on tumor cells by single and double stranded DNA cleavage after its activation, in which it blocks tumor cells’ DNA replication or transcription activities to inhibit tumor cells’ growth. Bleomycin sulfate (Blenoxane) is the most popular preparation used in clinical research, and contains Bleomycin fractions of A2 and B2, which causes difficulties in quantitative analysis. This work uses the metal chelating property of Bleomycin as an advantage to simplify and improve sensitivity of existing quantitative methods. Copper was spiked in excess to plasma samples, followed …
Il-17 Drives Copper Uptake And Activation Of Growth Pathways In Colorectal Cancer Cells In A Steap4-Dependent Manner, Evan Martin
Il-17 Drives Copper Uptake And Activation Of Growth Pathways In Colorectal Cancer Cells In A Steap4-Dependent Manner, Evan Martin
ETD Archive
Colorectal cancer is a disease characterized by abnormal, invasive cell growth beginning in the colon or rectum. The third most common type of cancer worldwide, approximately one million new cases of the disease are diagnosed across the globe annually, resulting in an estimated 700,000+ deaths. One major risk factor associated with development of colorectal cancer is the presence of chronic inflammation in the large intestine, also known as colitis. Inflammation is a complex immune response against harmful stimuli, characterized by symptoms including heat, redness, swelling and pain. One important molecular mediator of this process is interleukin 17 (IL-17), a pro-inflammatory …
The Future Perspective: Metabolomics In Laboratory Medicine For Inborn Errors Of Metabolism, Yana Sandlers
The Future Perspective: Metabolomics In Laboratory Medicine For Inborn Errors Of Metabolism, Yana Sandlers
Chemistry Faculty Publications
Metabolomics can be described as a simultaneous and comprehensive analysis of small molecules in a biological sample. Recent technological and bioinformatics advances have facilitated large-scale metabolomic studies in many areas, including inborn errors of metabolism (IEMs). Despite significant improvements in the diagnosis and treatment of some IEMs, it is still challenging to understand how genetic variation affects disease progression and susceptibility. In addition, a search for new more personalized therapies and a growing demand for tools to monitor the long-term metabolic effects of existing therapies set the stage for metabolomics integration in preclinical and clinical studies. While targeted metabolomics approach …
Development And Validation Of A Novel Lc–Ms/Ms Method For Simultaneous Determination Of Abiraterone And Its Seven Steroidal Metabolites In Human Serum: Innovation In Separation Of Diastereoisomers Without Use Of A Chiral Column, Mohammad Alyamani, Zhenfei Li, Sunil K. Upadhyay, David J. Anderson, Richard J. Auchus, Nima Sharifi
Development And Validation Of A Novel Lc–Ms/Ms Method For Simultaneous Determination Of Abiraterone And Its Seven Steroidal Metabolites In Human Serum: Innovation In Separation Of Diastereoisomers Without Use Of A Chiral Column, Mohammad Alyamani, Zhenfei Li, Sunil K. Upadhyay, David J. Anderson, Richard J. Auchus, Nima Sharifi
Chemistry Faculty Publications
Abiraterone acetate (AA), the prodrug of abiraterone, is FDA-approved for the treatment of castration-resistant prostate cancer. Abiraterone is metabolized in patients to a more potent analogue, D4A. However, we have recently reported that this analogue is further metabolized to additional metabolites in patients treated with AA. Here, we present a liquid chromatography-tandem mass spectrometry method developed to resolve and detect abiraterone and its seven metabolites in human serum using an AB Sciex Qtrap 5500 mass analyzer coupled with a Shimadzu Nexera UPLC station. Analytes and the internal standard (abiraterone-d4) were extracted from human serum using the liquid–liquid extraction procedure. The …
A Comparative Analysis Of Translesion Dna Synthesis Catalyzed By A High-Fidelity Dna Polymerase, Anvesh Dasari, Tejal Deodhar, Anthony J. Berdis
A Comparative Analysis Of Translesion Dna Synthesis Catalyzed By A High-Fidelity Dna Polymerase, Anvesh Dasari, Tejal Deodhar, Anthony J. Berdis
Chemistry Faculty Publications
Translesion DNA synthesis (TLS) is the ability of DNA polymerases to incorporate nucleotides opposite and beyond damaged DNA. TLS activity is an important risk factor for the initiation and progression of genetic diseases such as cancer. In this study, we evaluate the ability of a high-fidelity DNA polymerase to perform TLS with 8-oxo-guanine (8-oxo-G), a highly pro-mutagenic DNA lesion formed by reactive oxygen species. Results of kinetic studies monitoring the incorporation of modified nucleotide analogs demonstrate that the binding affinity of the incoming dNTP is controlled by the overall hydrophobicity of the nucleobase. However, the rate constant for the …
Inhibiting Translesion Dna Synthesis As An Approach To Combat Drug Resistance To Dna Damaging Agents, Jung-Suk Choi, Seol Kim, Edward Motea, Anthony J. Berdis
Inhibiting Translesion Dna Synthesis As An Approach To Combat Drug Resistance To Dna Damaging Agents, Jung-Suk Choi, Seol Kim, Edward Motea, Anthony J. Berdis
Chemistry Faculty Publications
Anti-cancer agents exert therapeutic effects by damaging DNA. Unfortunately, DNA polymerases can effectively replicate the formed DNA lesions to cause drug resistance and create more aggressive cancers. To understand this process at the cellular level, we developed an artificial nucleoside that visualizes the replication of damaged DNA to identify cells that acquire drug resistance through this mechanism. Visualization is achieved using "click" chemistry to covalently attach azide-containing fluorophores to the ethynyl group present on the nucleoside analog after its incorporation opposite damaged DNA. Flow cytometry and microscopy techniques demonstrate that the extent of nucleotide incorporation into genomic DNA is enhanced …
Hmba Is A Putative Hsp70 Activator Stimulating Hexim1 Expression That Is Down-Regulated By Estrogen, Rati Lama, Chunfang Gan, Nethrie Idippily, Viharika Bobba, David Danielpour, Monica Montano, Bin Su Ph.D.
Hmba Is A Putative Hsp70 Activator Stimulating Hexim1 Expression That Is Down-Regulated By Estrogen, Rati Lama, Chunfang Gan, Nethrie Idippily, Viharika Bobba, David Danielpour, Monica Montano, Bin Su Ph.D.
Chemistry Faculty Publications
Hexamethylene bis-acetamide inducible protein 1 (HEXIM1) is identified as a novel inhibitor of estrogen stimulated breast cell growth, and it suppresses estrogen receptor-a transcriptional activity. HEXIM1 protein level has been found to be downregulated by estrogens. Recently, HEXIM1 has been found to inhibit androgen receptor transcriptional activity as well. Researchers have used Hexamethylene bisacetamide (HMBA) for decades to stimulate HEXIM1 expression, which also inhibit estrogen stimulated breast cancer cell gene activation and androgen stimulated prostate cancer gene activation. However, the direct molecular targets of HMBA that modulate the induction of HEXIM1 expression in mammalian cells have not been identified. Based …
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
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 …
Metabolomics Reveals New Mechanisms For Pathogenesis In Barth Syndrome And Introduces Novel Roles For Cardiolipin In Cellular Function, Yana Sandlers, Kelly Mercier, Wimal Pathmasiri, Jim Carlson, Susan Mcritchie, Susan Sumner, Hilary J. Vernon
Metabolomics Reveals New Mechanisms For Pathogenesis In Barth Syndrome And Introduces Novel Roles For Cardiolipin In Cellular Function, Yana Sandlers, Kelly Mercier, Wimal Pathmasiri, Jim Carlson, Susan Mcritchie, Susan Sumner, Hilary J. Vernon
Chemistry Faculty Publications
Barth Syndrome is the only known Mendelian disorder of cardiolipin remodeling, with characteristic clinical features of cardiomyopathy, skeletal myopathy, and neutropenia. While the primary biochemical defects of reduced mature cardiolipin and increased monolysocardiolipin are well-described, much of the downstream biochemical dysregulation has not been uncovered, and biomarkers are limited. In order to further expand upon the knowledge of the biochemical abnormalities in Barth Syndrome, we analyzed metabolite profiles in plasma from a cohort of individuals with Barth Syndrome compared to age-matched controls via 1H nuclear magnetic resonance spectroscopy and liquid chromatography-mass spectrometry. A clear distinction between metabolite profiles of …
The Dual Regulatory Role Of Amino Acids Leu480 And Gln481 Of Prothrombin, Joesph R. Wiencek, Jamila Hirbawi, Vivien C. Yee, Michael Kalafatis
The Dual Regulatory Role Of Amino Acids Leu480 And Gln481 Of Prothrombin, Joesph R. Wiencek, Jamila Hirbawi, Vivien C. Yee, Michael Kalafatis
Chemistry Faculty Publications
Prothrombin (FII) is activated to α-thrombin (IIa) by prothrombinase. Prothrombinase is composed of a catalytic subunit, factor Xa (fXa), and a regulatory subunit, factor Va (fVa), assembled on a membrane surface in the presence of divalent metal ions. We constructed, expressed, and purified several mutated recombinant FII (rFII) molecules within the previously determined fVa-dependent binding site for fXa (amino acid region 473–487 of FII). rFII molecules bearing overlapping deletions within this significant region first established the minimal stretch of amino acids required for the fVa-dependent recognition exosite for fXa in prothrombinase within the amino acid sequence Ser478–Val479 …
Multi-Dimensional Glycan Microarrays With Glyco-Macroligands, Satya Nandana Narla, Huan Nie, Yu Li, Xue-Long Sun
Multi-Dimensional Glycan Microarrays With Glyco-Macroligands, Satya Nandana Narla, Huan Nie, Yu Li, Xue-Long Sun
Chemistry Faculty Publications
Glycan microarray has become a powerful high-throughput tool for examining binding interactions of carbohydrates with the carbohydrate binding biomolecules like proteins, enzymes, antibodies etc. It has shown great potential for biomedical research and applications, such as antibody detection and profiling, vaccine development, biomarker discovery, and drug screening. Most glycan microarrays were made with monovalent glycans immobilized directly onto the array surface via either covalent or non-covalent bond, which afford a multivalent glycans in two dimensional (2D) displaying. A variety of glyco-macroligands have been developed to mimic multivalent carbohydrate-protein interactions for studying carbohydrate-protein interactions and biomedical research and applications. Recently, a …
Cholesterol Conjugated Hdac Inhibitor As Novel Anticancer Agent, Paul Orefice, Jane Peterson, Bin Sun
Cholesterol Conjugated Hdac Inhibitor As Novel Anticancer Agent, Paul Orefice, Jane Peterson, Bin Sun
Undergraduate Research Posters 2014
Histone deacetylase (HDAC) inhibitors are a class of promising new multifunctional anticancer agents. These agents are able to affect multiple epigenetic changes in aberrant cells. In addition to regulating the gene expression and transcription via chromatin remodeling, HDAC inhibitors can also modulate a variety of cellular functions including proliferation, differentiation, and apoptosis. Vorinostat (SuberAniloHydroxamic Acid, SAHA), the first HDAC inhibitor approved by FDA, inhibited the metastasis of various cancer cells. However, SAHA distributes in cancer tissue and normal tissue in a similar level. It will be ideal to selectively delivery SAHA into cancer cells. Rapidly growing cancer cells have a …
Reclaiming The Efficacy Of Β-Lactam–Β-Lactamase Inhibitor Combinations: Avibactam Restores The Susceptibility Of Cmy-2-Producing Escherichia Coli To Ceftazidime, Krisztina M. Papp-Wallace, Marisa L. Winkler, Julian A. Gatta, Magdalena A. Taracila, Sujatha Chilakala, Yan Xu, J. Kristie Johnson, Robert A. Bonomo
Reclaiming The Efficacy Of Β-Lactam–Β-Lactamase Inhibitor Combinations: Avibactam Restores The Susceptibility Of Cmy-2-Producing Escherichia Coli To Ceftazidime, Krisztina M. Papp-Wallace, Marisa L. Winkler, Julian A. Gatta, Magdalena A. Taracila, Sujatha Chilakala, Yan Xu, J. Kristie Johnson, Robert A. Bonomo
Chemistry Faculty Publications
CMY-2 is a plasmid-encoded Ambler class C cephalosporinase that is widely disseminated in Enterobacteriaceae and is responsible for expanded-spectrum cephalosporin resistance. As a result of resistance to both ceftazidime and β-lactamase inhibitors in strains carrying blaCMY, novel β-lactam–β-lactamase inhibitor combinations are sought to combat this significant threat to β-lactam therapy. Avibactam is a bridged diazabicyclo [3.2.1]octanone non-β-lactam β-lactamase inhibitor in clinical development that reversibly inactivates serine β-lactamases. To define the spectrum of activity of ceftazidime-avibactam, we tested the susceptibilities of Escherichia coli clinical isolates that carry blaCMY-2 or blaCMY-69 and investigated the inactivation kinetics of CMY-2. Our analysis showed that …
Bsa–Boronic Acid Conjugate As Lectin Mimetics, Satya Nandana Narla, Poornima Pinnamaneni, Huan Nie, Yu Li, Xue-Long Sun
Bsa–Boronic Acid Conjugate As Lectin Mimetics, Satya Nandana Narla, Poornima Pinnamaneni, Huan Nie, Yu Li, Xue-Long Sun
Chemistry Faculty Publications
We report bovine serum albumin (BSA)–boronic acid (BA) conjugates as lectin mimetics and their glyco-capturing capacity. The BSA–BA conjugates were synthesized by amidation of carboxylic acid groups in BSA with aminophenyl boronic acid in the presence of EDC, and were characterized by Alizarin Red S (ARS) assay and SDS–PAGE gel. The BSA–BA conjugates were immobilized onto maleimide-functionalized silica beads and their sugar capturing capacity and specificity were confirmed by ARS displacement assay. Further, surface plasmon resonance (SPR) analysis of the glyco-capturing activity of the BSA–BA conjugates was conducted by immobilizing BSA–BA onto SPR gold chip. Overall, we demonstrated a BSA–BA-based …
Quantification Of Free Sialic Acid In Human Plasma Through A Robust Quinoxalinone Derivatization And Lc–Ms/Ms Using Isotope-Labeled Standard Calibration, Dan Wang, Xiang Zhou, Lin Wang, Sihe Wang, Xue-Long Sun
Quantification Of Free Sialic Acid In Human Plasma Through A Robust Quinoxalinone Derivatization And Lc–Ms/Ms Using Isotope-Labeled Standard Calibration, Dan Wang, Xiang Zhou, Lin Wang, Sihe Wang, Xue-Long Sun
Chemistry Faculty Publications
We report an accurate quantification of free sialic acid (SA) in human plasma using LC–MS/MS method with isotope-labeled standard calibration (ILSC) and robust derivatization. Specifically, derivatization of SA with a stable and inexpensive 3,4-diaminotoluene (DAT) provides a stable product of SA with high MS response, proving a convenient and cost-effective LC–MS/MS analysis of free SA. In addition, the use of 13C3-SA as calibration standard ensured the accuracy for the measurement. This assay used ultra high performance liquid chromatography (UHPLC) for separation of native/labeled SA and IS from matrix interference, and employed mass spectrometry in multiple reaction monitoring …
Combining Orthogonal Trna/Synthatase Pair And Amber Codon Suppression To Genetically Encode Oxidative Damage In High Density Lipoproteins, Jaclyn Alatrash, Nicholas Gilliam, Alisha House, Valentin Gogonea
Combining Orthogonal Trna/Synthatase Pair And Amber Codon Suppression To Genetically Encode Oxidative Damage In High Density Lipoproteins, Jaclyn Alatrash, Nicholas Gilliam, Alisha House, Valentin Gogonea
Undergraduate Research Posters 2013
Apolipoprotein A-I (apoA-I) is the main protein constituent of high density lipoprotein (HDL - the “good cholesterol”). Oxidatively damaged apoA-I has been isolated from circulating plasma and atherosclerosis plaque with the amino acid residue tryptophan 72 (W72) of apoA-I identified as a primary oxidation site. ApoA-I designed to include specific oxidized amino acids can be used to further investigate the role of site-specific oxidative damage in atherosclerosis. Genetic encoding of oxidized amino acids through orthogonal tRNA/aminoacyl-tRNA synthetase (aaRS) pairs offers a reliable method for producing site-specific oxidized proteins. Our project involves the generation of Saccharomyces tryptophan-RS mutants for recognition of …
Insights Into The Roles Of Desolvation And Π-Electron Interactions During Dna Polymerization, Edward A. Motea, Irene Lee, Anthony J. Berdis
Insights Into The Roles Of Desolvation And Π-Electron Interactions During Dna Polymerization, Edward A. Motea, Irene Lee, Anthony J. Berdis
Chemistry Faculty Publications
This report describes the use of several isosteric non-natural nucleotides as probes to evaluate the roles of nucleobase shape, size, solvation energies, and π-electron interactions as forces influencing key kinetic steps of the DNA polymerization cycle. Results are provided using representative high- and low-fidelity DNA polymerases. Results generated with the E. coli Klenow fragment reveal that this high-fidelity polymerase utilizes hydrophobic nucleotide analogues with higher catalytic efficiencies compared to hydrophilic analogues. These data support a major role for nucleobase desolvation during nucleotide selection and insertion. In contrast, the low-fidelity HIV-1 reverse transcriptase discriminates against hydrophobic analogues and only tolerates non-natural …
Spectroscopic Analysis Of Polymerization And Exonuclease Proofreading By A High-Fidelity Dna Polymerase During Translesion Dna Synthesis, Babho Devadoss, Irene Lee, Anthony J. Berdis
Spectroscopic Analysis Of Polymerization And Exonuclease Proofreading By A High-Fidelity Dna Polymerase During Translesion Dna Synthesis, Babho Devadoss, Irene Lee, Anthony J. Berdis
Chemistry Faculty Publications
High fidelity DNA polymerases maintain genomic fidelity through a series of kinetic steps that include nucleotide binding, conformational changes, phosphoryl transfer, polymerase translocation, and nucleotide excision. Developing a comprehensive understanding of how these steps are coordinated during correct and pro-mutagenic DNA synthesis has been hindered due to lack of spectroscopic nucleotides that function as efficient polymerase substrates. This report describes the application of a non-natural nucleotide designated 5-naphthyl-indole-2′-deoxyribose triphosphate which behaves as a fluorogenic substrate to monitor nucleotide incorporation and excision during the replication of normal DNA versus two distinct DNA lesions (cyclobutane thymine dimer and an abasic site). Transient …
Voltage-Controlled Enzyme-Catalyzed Glucose–Gluconolactone Conversion Using A Field-Effect Enzymatic Detector, Siu Tung Yau, Yan Xu, Yang Song, Ye Feng, Jiapeng Wang
Voltage-Controlled Enzyme-Catalyzed Glucose–Gluconolactone Conversion Using A Field-Effect Enzymatic Detector, Siu Tung Yau, Yan Xu, Yang Song, Ye Feng, Jiapeng Wang
Chemistry Faculty Publications
The field-effect enzymatic detection (FEED) technique was used to control the kinetics of the enzymatic conversion of glucose to gluconolactone. The glucose–gluconolactone conversion occurring at an enzyme-immobilized electrode, a well-studied process, was confirmed using mass spectrometry. Electrochemical studies showed that the glucose oxidation current depends on the gating voltage VG and the ion concentration of the sample solution. Additionally, the depletion of glucose in the sample also showed a dependence on VG. FEED was used to detect H2O2 on the zepto-molar level in order to show the ultrasensitive detection capability of the technique. These results, while providing evidence for the …
Exploring The Role Of A Conserved Class A Residue In The Ω-Loop Of Kpc-2 Β-Lactamase: A Mechanism For Ceftazidime Hydrolysis
Chemistry Faculty Publications
Gram-negative bacteria harboring KPC-2, a class A β-lactamase, are resistant to all β-lactam antibiotics and pose a major public health threat. Arg-164 is a conserved residue in all class A β-lactamases and is located in the solvent-exposed Ω-loop of KPC-2. To probe the role of this amino acid in KPC-2, we performed site-saturation mutagenesis. When compared with wild type, 11 of 19 variants at position Arg-164 in KPC-2 conferred increased resistance to the oxyimino-cephalosporin, ceftazidime (minimum inhibitory concentration; 32→128 mg/liter) when expressed in Escherichia coli. Using the R164S variant of KPC-2 as a representative β-lactamase for more detailed analysis, we …
Understanding The Molecular Determinants Of Substrate And Inhibitor Specificities In The Carbapenemase Kpc-2: Exploring The Roles Of Arg220 And Glu276
Chemistry Faculty Publications
β-Lactamases are important antibiotic resistance determinants expressed by bacteria. By studying the mechanistic properties of β-lactamases, we can identify opportunities to circumvent resistance through the design of novel inhibitors. Comparative amino acid sequence analysis of class A β-lactamases reveals that many enzymes possess a localized positively charged residue (e.g., R220, R244, or R276) that is critical for interactions with β-lactams and β-lactamase inhibitors. To better understand the contribution of these residues to the catalytic process, we explored the roles of R220 and E276 in KPC-2, a class A β-lactamase that inactivates carbapenems and β-lactamase inhibitors. Our study reveals that substitutions …