Biochemistry, Biophysics, and Structural Biology Commons^™

Fabrication Of Polymeric Microfluidic Devices For Protein Analysis, Jikun Liu

Theses and Dissertations

2-Bromoisobutyryl bromide was immobilized on poly(methyl methacrylate) (PMMA) substrates activated using an oxygen plasma. Atom-transfer radical polymerization was then performed to graft poly(ethylene glycol) (PEG) on the PMMA surface. PMMA micro capillary electrophoresis (µCE) devices made with the covalently modified surfaces exhibited substantially reduced electroosmotic flow and nonspecific adsorption of proteins. Both column efficiency and migration time reproducibility were one order of magnitude better with derivatized PMMA µCE devices compared to untreated versions. Fast, reproducible, and efficient separations of proteins and peptides were demonstrated using the PEG-grafted PMMA µCE chips. All analyses were completed in less than 60 seconds, and …

(Review) Green Fluorescent Proteins, Marc Zimmer

Chemistry Faculty Publications

Reviews the book:Green Fluorescent Protein: Properties, Applications, and Protocols. Second Edition. Methods of Biochemical Analysis, Volume 47. Edited by Martin Chalfie and Steven R Kain.Green Fluorescent Protein: Properties, Applications, and Protocols. Second Edition. Methods of Biochemical Analysis, Volume 47. Edited by Martin Chalfie and Steven R Kain. Hoboken (New Jersey): Wiley-Interscience. $89.95. xv + 443 p + 24 pl; ill.; index. ISBN: 0–471–73682–1. 2006.

Mechanisms Imposing The Vbeta Bias Of V14a Natural Killer T Cells And Consequences For Microbial Glycolipid Recognition, Paul B. Savage, Dasten G. Wei, Shane A. Curran, Luc Teyton, Albert Bendelac

Faculty Publications

Mouse and human natural killer T (NKT) cells recognize a restricted set of glycosphingolipids presented by CD1d molecules, including self iGb3 and microbial alpha-glycuronosylceramides. The importance of the canonical Valpha 14-Jalpha18 TCR alphachain for antigen recognition by NKT cells is well recognized, but the mechanisms underlying the Vbeta8, Vbeta7, and Vbeta2 bias in mouse have not been explored. To study the influences of thymic selection and the constraints of pairing with Valpha 14-Jalpha 18, we have created a population of mature T cells expressing Valpha 14-Jalpha 18 TCRalpha chain in CD1d-deficient mice and studied its recognition properties in vitro and …

Biogeochemistry Of Arsenic And Antimony In The North Pacific Ocean, Gregory A. Cutter, Lynda S. Cutter

OES Faculty Publications

The biogeochemical cycles of the metalloid elements arsenic and antimony were examined along a 15,000 km surface water transect and at 9 vertical profile stations in the western North Pacific Ocean as part of the 2002 IOC Contaminant Baseline Survey. Results show that the speciation of dissolved arsenic (As III, As V, and methylated As) was subtly controlled by the arsenate (AsV)/phosphate ratio. An additional fraction of presumed organic arsenic previously reported in coastal waters was also present (~15% of the total As) in oceanic surface waters. Dissolved inorganic antimony displayed mildly scavenged behavior that was confirmed by correlations with …

Erratum: "Hard Sphere Radial Distribution Function Again", Andrij Trokhymchuk, Douglas Henderson, Ivo Nezbeda, Jan Jirsak

Faculty Publications

Some misprints have been found for parameters given by Eqs. 29 and 30 and the relevant equations in the Appendix.

Two New Resorcinarenes: A Pyridine & Acetic Acid Ligand For Metal Coordination And A Deep-Cavity Nitroquinoxaline Resorcinarene, Samantha Sizemore Vernetti

Theses and Dissertations

Functionalizing the upper rim of resorcinarene-based cavitands allows a variety of compounds to be synthesized from a single scaffold. Using the upper-rim moieties as ligands for a variety of transition metal ions further increases the versatility of this class of host compounds. A new resorcinarene-based molecule functionalized with four pyridine and acetic acid ligands has been successfully prepared to explore the properties of metal-assembled complexes. To synthesize this compound, tetra(bromomethyl)cavitand was reacted with N-(2-pyridylmethyl)-N-ethylacetate amine to give ethyl acetate pyridine resorcinarene. Hydrolysis of the ester gave acetic acid pyridine resorcinarene (APRes) in good yield. Complexes with Cu2+, Co2+, and Zn2+ …

Development Of A High Spatial Selectivity Tri-Polar Concentric Ring Electrode For Laplacian Electroencephalography (Leeg) System, Kanthaiah Koka

Doctoral Dissertations

Brain activity generates electrical potentials that are spatio-temporal in nature. Electroencephalography (EEG) is the least costly and most widely used non-invasive technique for diagnosing many brain problems. It has high temporal resolution but lacks high spatial resolution.

The surface Laplacian will enhance the spatial resolution of EEG as it performs the second spatial derivative of the surface potentials. In an attempt to increase the spatial selectivity, researchers introduced a bipolar electrode configuration using a five point finite difference method (FPM) and others applied a quasi-bipolar (tri-polar with two elements shorted) concentric electrode configuration. To further increase the spatial resolution, the …

Gas Chromatography: Mass Spectrometry Of Chemical Agents And Related Interferents, Lailiang Zhai

Theses and Dissertations

One of the main problems encountered in chemical analysis operations in the field is collecting sufficient sample from the source and transferring that sample to the measurement instrument for fast separation and identification. I have been involved in developing a field-portable gas chromatography-mass spectrometry (GC-MS) system with solid phase microextraction (SPME) sampling for point detection of chemical agents. The objective is to minimize the analysis time between sampling and detection of a potential chemical threat. SPME offers a convenient means for sampling gaseous and liquid samples, concentrating the analytes, and transferring the analytes to the injection port of a GC …

Development Of An Unbiased Statistical Method For The Analysis Of Unigenic Evolution, Colleen D. Behrsin, Chris J. Brandl, David W. Litchfield, Brian H. Shilton, Lindi M. Wahl

Biochemistry Publications

Background: Unigenic evolution is a powerful genetic strategy involving random mutagenesis of a single gene product to delineate functionally important domains of a protein. This method involves selection of variants of the protein which retain function, followed by statistical analysis comparing expected and observed mutation frequencies of each residue. Resultant mutability indices for each residue are averaged across a specified window of codons to identify hypomutable regions of the protein. As originally described, the effect of changes to the length of this averaging window was not fully eludicated. In addition, it was unclear when sufficient functional variants had been examined …

Membrane-Based Protein Preconcentration Microfluidic Devices, Yi Li

Theses and Dissertations

Interest in microchip capillary electrophoresis (CE) is growing due to the rapid analysis times provided and small sample input requirements. However, higher-concentration samples are typically needed because of the small (~pL) detection volumes in these devices. I have made membrane-based protein preconcentration systems in capillary and microchip designs to increase the detectability of low-concentration biological samples. A photopolymerized ion-permeable membrane interfaced with a microchannel in poly(methyl methacrylate) (PMMA) formed the preconcentrator. When a voltage was applied between the sample reservoir and the ionically conductive membrane in a capillary-based system, R-phycoerythrin was concentrated more than 1,000 fold, as determined by laser-induced …

Hydrographic Observations During The 2002 Ioc Contaminant Baseline Survey In The Western Pacific Ocean, C. I. Measures, Greg A. Cutter, W. M. Landing, R. T. Powell

OES Faculty Publications

The 2002 IOC Contaminant Baseline Survey in the western Pacific Ocean was the fourth in a series of cruises intended to establish the contemporary concentrations of trace elements and other materials in the major water masses of the ocean and to illuminate the pathways by which materials delivered to the surface ocean are incorporated in the subsurface waters. The expedition occupied 9 vertical profile stations encompassing the subtropical and subarctic gyre of the western North Pacific. In addition, underway surface water samples were collected during transits between the stations. This paper uses the temperature, salinity, nutrient, oxygen, and chlorophyll data …

Measurement, Characterization, And Source Apportionment Of The Major Chemical Components Of Fine Particulate Material, Including Semi-Volatile Species, Brett D. Grover

Theses and Dissertations

The promulgation of revised standards for atmospheric fine particles (PM2.5) by the US EPA has sparked renewed interest in the ability to accurately measure and characterize suspended atmospheric particulate matter. Semi-volatile material (SVM), consisting of ammonium nitrate and semi-volatile organic material (SVOM), is not accurately measured by EPA accepted methods such as the Federal reference method (FRM) or Tapered Element Oscillating Microbalance (TEOM). However, SVM is often a major fraction of urban aerosols. Recent advances in atmospheric sampling instrumentation allowed for the semi-continuous characterization of urban PM2.5, including SVM. The Filter Dynamic Measurement System (FDMS) was shown to measure total …

Characterization, Functionalization And Applications Of Alkyl Monolayers On Silicon Surfaces, Guilin Jiang

Theses and Dissertations

Investigations were performed on the stability, mechanism of formation and an application of alkyl monolayers chemomechanically prepared on silicon surfaces. A new method of surface modification, laser-activation modification of surfaces (LAMS), and multivariate analyses of time-of-flight secondary ion mass spectrometry (ToF-SIMS) images of LAMS spots were also reported. X-ray photoelectron spectroscopy (XPS) and other data show that alkyl monolayers prepared by scribing silicon under 1-iodoalkanes and 1-alkenes were stable over extended periods of time to air, water, a boiling acid and Al Ka X-rays. The stability is attributed to direct Si-C bonding in the monolayers. The observation that the oxygen …

Exchange Coupling And Contribution Of Induced Orbital Angular Momentum Of Low-Spin Fe^3+ Ions To Magnetic Anisotropy In Cyanide-Bridged Fe2m2 Molecular Magnets: Spin-Polarized Density-Functional Calculations, Stephen Holmes, Kyungwha Park

Chemistry & Biochemistry Faculty Works

Electronic structure and intramolecular exchange constants are calculated for three cyanide-bridged molecular magnets, [Tp⋆FeIII(CN)3MII(DMF)4]2(OTf)2∙2DMF (MII=Mn,Co,Ni) (abbreviated as Fe2Mn2, Fe2Co2, and Fe2Ni2) that have been recently synthesized, within a generalized-gradient approximation in spin-polarized density-functional theory (DFT). Here Tp⋆=[C3(CH3)2HN2]3BH, OTf=O3SCF3, and DMF=HCON(CH3)2. Due to strong ligand fields present in the [Tp⋆FeIII(CN)3]− units, the Fe3+ ions exhibit a low ground-state spin of S=1/2. Our calculations show that the metal ions in the Fe2Mn2 molecule interact antiferromagnetically via cyanide ligands, while those in the Fe2Co2 and Fe2Ni2 molecule interact ferromagnetically. The calculations also suggest that the smallest gaps between the highest occupied molecular orbital …

Fluorescent Analysis Of Translesion Dna Synthesis By Using A Novel, Non-Natural Nucleotide Analogue, Irene Lee, Anthony J. Berdis

Chemistry Faculty Publications

The replication of damaged DNA is a promutagenic process that can lead to disease development. This report evaluates the dynamics of nucleotide incorporation opposite an abasic site, a commonly formed DNA lesion, by using two fluorescent nucleotide analogues, 2-aminopurine deoxyribose triphosphate (2-APTP) and 5-phenylindole deoxyribose triphosphate (5-PhITP). In both cases, the kinetics of incorporation were compared by using a 32 P-radiolabel extension assay versus a fluorescence-quenching assay. Although 2-APTP is efficiently incorporated opposite a templating nucleobase (thymine), the kinetics for incorporation opposite an abasic site are significantly slower. The lower catalytic efficiency hinders its use as a probe to study …

Recent Developments Inthe Mechanistic Enzymology Of The Atp-Dependent Lon Protease From Escherichia Coli: Highlights From Kinetic Studies, Irene Lee, Anthony J. Berdis, Carolyn K. Suzuki

Chemistry Faculty Publications

Lon protease, also known as protease La, is one of the simplest ATP-dependent proteases that plays vital roles in maintaining cellular functions by selectively eliminating misfolded, damaged and certain short-lived regulatory proteins. Although Lon is a homo-oligomer, each subunit of Lon contains both an ATPase and a protease active site. This relatively simple architecture compared to other hetero-oligomeric ATP-dependent proteases such as the proteasome makes Lon a useful paradigm for studying the mechanism of ATP-dependent proteolysis. In this article, we survey some recent developments in the mechanistic characterization of Lon with an emphasis on the utilization of pre-steady-state enzyme kinetic …

Structural And Thermodynamic Parameters Associated With The Interaction Of Cobalt (Iii) Complexes And Duplex Dna, Jaime M. Ferreira

Seton Hall University Dissertations and Theses (ETDs)

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Potential Of Vibrational Spectroscopy In The Diagnosis Of Human Tumours., Eoghan O'Faolain

Doctoral

Just fewer than 20,000 people are annually diagnosed with some form of cancer in Ireland and one in three people are likely to contract some form of cancer by age 74. With the number of cases increasing at an annual rate of 2%, the early detection and treatment of cancer is becoming increasingly important. Both IR and Raman spectroscopy offer the potential for real time, quantitative detection of cancer and even precancer. This study investigates the potential of Raman and Fourier transform infrared, both benchtop and synchrotron spectroscopies for the detection of cervical cancer. The tissue was classified and its …

Dna Damage Responses In Progeroid Syndromes Arise From Defective Maturation Of Prelamin A, Michael Sinensky, Y. Liu, A. Rusinol, Y. Wang, Y. Zou

Faculty Publications, Biological Sciences

The genetic diseases Hutchinson-Gilford progeria syndrome (HGPS) and restrictive dermopathy (RD) arise from accumulation of farnesylated prelamin A because of defects in the lamin A maturation pathway. Both of these diseases exhibit symptoms that can be viewed as accelerated aging. The mechanism by which accumulation of farnesylated prelamin A leads to these accelerated aging phenotypes is not understood. Here we present evidence that in HGPS and RD fibroblasts, DNA damage checkpoints are persistently activated because of the compromise in genomic integrity. Inactivation of checkpoint kinases Ataxia-telangiectasia-mutated (ATM) and ATR (ATM- and Rad3-related) in these patient cells can partially overcome their …

Farnesylated Lamins, Progeroid Syndromes And Farnesyl Transferase Inhibitors, Michael Sinensky, A. E. Rusinol

Faculty Publications, Biological Sciences

Three mammalian nuclear lamin proteins, lamin B1, lamin B2 and the lamin A precursor, prelamin A, undergo canonical farnesylation and processing at CAAX motifs. In the case of prelamin A, there is an additional farnesylation-dependent endoproteolysis, which is defective in two congenital diseases: Hutchinson-Gilford progeria (HGPS) and restrictive dermopathy (RD). These two diseases arise respectively from defects in the prelamin A substrate and the enzyme (ZmpSte24) that processes it. Recent work has shed light on the roles of the lamin proteins and the enzymes involved in their farnesylation-dependent maturation. Other experimental work, including mouse model studies, have examined the possibility …

Process For The Preparation Of Citalopram Intermediate, Aslam A. Malik, Hasan Palandoken, Joy A. Stringer, Dershing Huang, Antonio Romero, Olivier Dapremont

Chemistry and Biochemistry

The present invention provides, inter alia, a novel process for the preparation of Citalopram, a known antidepressant.

The Role Of Phosducin-Like Protein And The Cytosolic Chaperonin Cct In G Beta Gamma Dimer Assembly, Ting Hu

Theses and Dissertations

Phosducin-like protein (PhLP), a G protein beta gamma subunit dimer binder and G protein signaling regulator, was suggested to regulate the activity of cytosolic chaperonin CCT by their high affinity interaction. In the present study, the three-dimensional structure of PhLP:CCT complex has been solved by cryoelectron microscopy. PhLP was found to bind only one of the chaperonin rings with both N- and C-terminal domains. It spans the central folding cavity of CCT and interacts with two opposite sides of the top apical region, inducing the constraining of the entry of the folding cavity. These findings support a putative role of …

Palladium-Imidazolium Carbene Catalyzed Heck Coupling Reactions And Synthesis Of A Novel Class Of Fluoroanthracenylmethyl Ptc Catalysts, Jiuqing Zhang

Theses and Dissertations

Palladium catalyzed Heck coupling with aryl and alkenyl halides has become a powerful means of carbon-carbon bond formation. This standard synthetic method has been developed to a high level of utility using various catalysts, conditions and substrates. Yet significant drawbacks remain, including poor reactivity, the need for high temperatures and base, limited substrate generality, and selectivity. Mixed products often suffer from olefin migration following insertion. N-Heterocyclic carbenes (NHC) have proven to be electron-rich donors which provide higher stability and reactivity than phosphines. In a previous paper reported by our research group the imidazolium-palladium carbene has proven to be highly efficient …

Performance Of Binary-Encounter-Bethe (Beb) Theory For Electron-Impact Ionization Cross Sections Of Molecules Containing Heavy Elements (Z > 10), Gregory E. Scott, Karl K. Irikura

Chemistry and Biochemistry

The binary-encounter-Bethe (BEB) theory developed by Kim and coworkers has been successful for computing electron-impact ionization cross sections of many molecules. However, some recent publications have stated that BEB theory performs poorly for molecules that contain heavier elements such as chlorine and sulfur. We have found that the BEB calculations in those publications were performed incorrectly. When performed correctly, BEB predictions are as good for heavy-element molecules as for light-element molecules. We recommended recently that an alternative, less-confusing procedure be used for molecules that contain heavier elements. The alternative procedure, based upon effective core potentials (ECPs), does not require explicit …

Total Syntheses Of (+)-Geldanamycin, (-)-Ragaglitazar, And (+)-Kurasoin A And Phase-Transfer-Catalyzed Asymmetric Alkylation, Erik J. Hicken

Theses and Dissertations

Geldanamycin possesses various biological activities as seen in the NCI 60 cell line panel (13 nM avg., 70 nM SKBr-3 cells). The predominant mode of action providing these unique results arises from the ability of geldanamycin (GA) to bind to the chaperone heat shock protein 90 (Hsp90). Despite its complicated functionality, the first total synthesis of GA was accomplished, which included two new reactions developed specifically to address the stereochemical features. The final step in the synthesis of GA was a demethylation-oxidation sequence to generate the desired para-quinone. This step could only be accomplished with HNO3/AcOH, producing GA in …

Specific Heat And Thermodynamic Properties Of Metallic Systems: Instrumentation And Analysis, Brian E. Lang

Theses and Dissertations

A small-scale adiabatic calorimeter has been constructed as part of a larger project to study nano-particles and to facilitate specific heat measurements on samples where it is difficult to obtain enough material to run on the current large-scale adiabatic apparatus. This calorimeter is designed to measure sample sizes of less than 0.8 cc over a temperature range from 13 K to 350 K. Specific heat results on copper, sapphire, and benzoic acid show the accuracy of the measurements to be better than ±0.4% for temperatures higher than 50 K. The reproducibility of these measurements is generally better than ±0.25%. Experimental …

Microcapsule Biosensors Based On Competitive Binding And Fluorescence Resonance Energy Transfer Assays, Swetha Chinnayelka

Doctoral Dissertations

Fluorescent sensing systems offer the potential for minimally invasive monitoring with implantable devices, but they require carrier technologies that provide suitable immobilization, accessibility, and biocompatibility while maintaining adequate response characteristics. Towards the development of this goal, a general design of a biosensor with the capability of detecting different metabolites was investigated. The approach is based on the encapsulation of a competitive binding assay in microcapsules and monitoring the changes in fluorescence resonance energy transfer (FRET) in the presence of analyte. To experimentally demonstrate this type of sensing system, glucose was chosen as the model target analyte. The design, fabrication, and …

A Facile Synthesis Of (Tert-Alkoxy)Amines, Hasan Palandoken, Chris M. Bocian, Michelle R. Mccombs, Michael H. Nantz

Chemistry and Biochemistry

Tertiary alcohols react with stoichiometric BF₃·Et₂O and N-hydroxyphthalimide to yield N-alkoxyphthalimides. Subsequent hydrazinolyses afford the title compounds.

Polymer Microchips For Capillary Electrophoresis And Electric Field Gradient Focusing Of Biomolecules, Ryan Thomas Kelly

Theses and Dissertations

Polymeric materials have seen increasing use as microfluidic device substrates due to their low cost and the simplicity of templated fabrication procedures. I showed that poly(methyl methacrylate) (PMMA) microdevices could be enclosed in a boiling water bath, which allowed the seal to form more quickly than in conventional approaches, and enabled microchannels to remain hydrated throughout the bonding process. Microchip capillary electrophoresis (µ-CE) devices were fabricated using water-based enclosure, and a mixture of fluorescently labeled amino acids was separated in 30 s in these microchips. To create more robust capillary electrophoresis (CE) microdevices with improved separation performance, phase-changing sacrificial materials …

Structure Elucidation Of Bioactive Compounds Isolated From Endophytes Of Alstonia Scholaris And Acmena Graveolens, Nicholas James Hundley

Theses and Dissertations

Alstonia scholaris is an evergreen tree native to Southeast Asia and Australia. It is commonly used as a medicinal plant throughout these regions. In the present study, an endophyte of the genus Xylaria was isolated from a stem of Alstonia scholaris, its mycelia and exudate extracted, and the extract assayed for growth inhibition of HeLa cancer cells in vitro. Several known compounds were isolated and identified based on NMR, infrared, and mass spectral data. The compounds identified are 19,20-epoxycytochalasin C; 19,20epoxycytochalasin D; and xylobovide. Two other compounds, fusaric acid and dehydrofusaric acid, were discovered in an endophyte of the Hypocreales …