Biochemistry Commons^™

A Natural Missing Link Between Activated And Downhill Protein Folding Scenarios, Feng Liu, Caroline Maynard, Gregory E. Scott, Artem Melnykov, Kathleen B. Hall, Martin Gruebele

Chemistry and Biochemistry

We propose protein PTB1:4W as a good candidate for engineering into a downhill folder. PTB1:4W has a probe-dependent thermal unfolding curve and sub-millisecond T-jump relaxation kinetics on more than one time scale. Its refolding rate in denaturant is a non-linear function of denaturant concentration (curved chevron plot). Yet at high denaturant concentration its unfolding is probe-independent, and the folding kinetics can be fitted to a single exponential decay. The domain appears to fold via a mechanism between downhill folding and activated folding over several small barriers, and when denaturant is added, one of these barriers greatly increases and simplifies the …

Very-Long-Chain Fatty Acids Are Involved In Polar Auxin Transport And Developmental Patterning In Arabidopsis, Françoise Roudier, Lionel Gissot, Frédéric Beaudoin, Richard Haslam, Louise V. Michaelson, Jessica Marion, Diana Molino, Amparo Lima, Liên Bach, Halima Morin, Frédérique Tellier, Jean-Christophe Palauqui, Yannick Bellec, Charlotte Renne, Martine Miquel, Marco Dacosta, Julien Vignard, Christine Rochat, Jennifer E. Markham, Patrick Moreau, Jonathan Napier, Jean-Denis Faure

Department of Biochemistry: Faculty Publications

Very-long-chain fatty acids (VLCFAs) are essential for many aspects of plant development and necessary for the synthesis of seed storage triacylglycerols, epicuticular waxes, and sphingolipids. Identification of the acetyl-CoA carboxylase PASTICCINO3 and the 3-hydroxy acyl-CoA dehydratase PASTICCINO2 revealed that VLCFAs are important for cell proliferation and tissue patterning. Here, we show that the immunophilin PASTICCINO1 (PAS1) is also required for VLCFA synthesis. Impairment of PAS1 function results in reduction of VLCFA levels that particularly affects the composition of sphingolipids, known to be important for cell polarity in animals. Moreover, PAS1 associates with several enzymes of …

Nmr Assignments Of A Stable Processing Intermediate Of Human Frataxin, Kalyan C. Kondapalli, Krisztina Z. Bencze, Eric Dizin, James A. Cowan, Timothy L. Stemmler

Biochemistry and Molecular Biology Faculty Publications

Frataxin, a nuclear encoded protein targeted to the mitochondrial matrix, has recently been implicated as an iron chaperone that delivers ferrous iron to the iron-sulfur assembly enzyme IscU. During transport across the mitochondrial membrane, the N-terminal mitochondrial targeting sequence of frataxin is cleaved in a two-step process to produce the mature protein found in the matrix, however N-terminal extended forms of the protein have also been observed in vivo. The recent structural characterization studies of the human frataxin ortholog were performed on a truncated variant of the protein. Here we report the NMR spectral assignment of an extended form of …

Antispasmodic And Vasodilator Activities Of Morinda Citrifolia Root Extract Are Mediated Through Blockade Of Voltage Dependent Calcium Channels, Anwarul Hassan Gilani, Saf-Ur-Rehman Mandukhail, Javeid Iqbal, Masoom Yasinzai, Nauman Aziz, Aslam Khan, Najeeb-Ur-Rehman

Department of Biological & Biomedical Sciences

Background: Morinda citrifolia (Noni) is an edible plant with wide range of medicinal uses. It occurs exclusively in tropical climate zone from India through Southeast Asia and Australia to Eastern Polynesia and Hawaii. The objective of this study was to explore the possible mode(s) of action for its antispasmodic, vasodilator and cardio-suppressant effects to rationalize its medicinal use in gut and cardiovascular disorders. Methods: Isolated tissue preparations such as, rabbit jejunum, rat and rabbit aorta and guinea pig atria were used to test the antispasmodic and cardiovascular relaxant effects and the possible mode of action(s) of the 70% aqueous-ethanolic extract …

Spermine Oxidase (Smo) Activity In Breast Tumor Tissues And Biochemical Analysis Of The Anticancer Spermine Analogues Benspm And Cpenspm, Manuela Cervelli, Gabriella Bellavia, Emiliano Fratini, Roberto Amendola, Fabio Polticelli, Marco Barba, Rodolfo Federico, Fabrizio Signore, Giacomo Gucciardo, Rosalba Grillo, Patrick M. Woster, Robert A. Casero Jr, Paolo Mariottini

Wayne State University Associated BioMed Central Scholarship

Abstract

Background

Polyamine metabolism has a critical role in cell death and proliferation representing a potential target for intervention in breast cancer (BC). This study investigates the expression of spermine oxidase (SMO) and its prognostic significance in BC. Biochemical analysis of Spm analogues BENSpm and CPENSpm, utilized in anticancer therapy, was also carried out to test their property in silico and in vitro on the recombinant SMO enzyme.

Methods

BC tissue samples were analyzed for SMO transcript level and SMO activity. Student's t test was applied to evaluate the significance of the differences in value observed in T and NT …

Determining The Rate Of Transcription Of T7 Rna Polymerase Using Single Molecule Fluorescence Imaging, Dawn Renee Nichola

Theses, Dissertations and Capstones

It is important to understand the many factors impacting the rate at which an RNA polymerase incorporates nucleotides. The transcription rate of T7 RNA polymerase has been determined using single molecule fluorescence microscopy. A Cy3 labeled circular 45nt ssDNA molecule was used to monitor the transcription process. T7 RNA polymerase was used because it is a single subunit polymerase that does not need any cofactors and will transcribe single-stranded DNA circles that do not contain a promoter. The transcription was monitored by measuring the quasi-periodic change in intensity associated with the transit of the probe through the polymerase as the …

Electrochemical Sensing In Paper-Based Microfluidic Devices, Zhihong Nie, Christian A. Nijhuis, Jinlong Gong, Xin Chen, Alexander Kumachev, Andres W. Martinez, Max Narovlyansky, George M. Whitesides

Chemistry and Biochemistry

This paper describes the fabrication and the performance of microfluidic paper-based electrochemical sensing devices (we call the microfluidic paper-based electrochemical devices, μPEDs). The μPEDs comprise paper-based microfluidic channels patterned by photolithography or wax printing, and electrodes screen-printed from conducting inks (e.g., carbon or Ag/AgCl). We demonstrated that the μPEDs are capable of quantifying the concentrations of various analytes (e.g., heavy-metal ions and glucose) in aqueous solutions. This low-cost analytical device should be useful for applications in public health, environmental monitoring, and the developing world.

Mechanistic Importance Of Redox Potentials And Conformational Flexibility In Electron-Transferring Flavoproteins, Michael Anthony Swanson

Electronic Theses and Dissertations

The mitochondrial matrix flavoproteins electron transfer flavoprotein (ETF) and electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO) are responsible for linking fatty acid β-oxidation with the main mitochondrial respiratory chain. Electrons derived from flavoprotein dehydrogenases are transferred sequentially through ETF and ETF-QO to ubiquinone and then into the respiratory chain via complex III. In this study, the effects of changes in ETF-QO redox potentials on its activity and the conformational flexibility of ETF were investigated.

ETF-QO contains one [4Fe-4S]^2+,1+ and one flavin adenine dinucleotide (FAD). In the porcine protein, threonine 367 is hydrogen bonded to N1 and O2 of the flavin ring …

Programmable Diagnostic Devices Made From Paper And Tape, Andres W. Martinez, Scott T. Phillips, Zhihong Nie, Chao-Min Cheng, Emanuel Carrilho, Benjamin J. Wiley, George M. Whitesides

Chemistry and Biochemistry

This paper describes three-dimensional microfluidic paper-based analytical devices (3-D μPADs) that can be programmed (postfabrication) by the user to generate multiple patterns of flow through them. These devices are programmed by pressing single-use ‘on’ buttons, using a stylus or a ballpoint pen. Pressing a button closes a small space (gap) between two vertically aligned microfluidic channels, and allows fluids to wick from one channel to the other. These devices are simple to fabricate, and are made entirely out of paper and double-sided adhesive tape. Programmable devices expand the capabilities of μPADs and provide a simple method for controlling the movement …

Millimeter-Scale Contact Printing Of Aqueous Solutions Using A Stamp Made Out Of Paper And Tape, Chao-Min Cheng, Aaron D. Mazzeo, Jinlong Gong, Andres W. Martinez, Scott T. Phillips, Nina Jain, George M. Whitesides

Chemistry and Biochemistry

This communication describes a simple method for printing aqueous solutions with millimeter-scale patterns on a variety of substrates using an easily fabricated, paper-based microfluidic device (a paper-based ―stamp‖) as a contact printing device. The device is made from inexpensive materials, and it is easily assembled by hand; this method is thus accessible to a wide range of laboratories and budgets. A single device was used to print over 2500 spots in less than three minutes at a density of 16 spots per square centimetre. This method provides a new tool to pattern biochemicals—reagents, antigens, proteins, and DNA—on planar substrates. The …

Trace Gas And Particle Emissions From Domestic And Industrial Biofuel Use And Garbage Burning In Central Mexico, Ted J. Christian, Robert Yokelson, B. Cardenas, L. T. Molina, G. Engling, S. C. Hsu

Chemistry and Biochemistry Faculty Publications

In central Mexico during the spring of 2007 we measured the initial emissions of 12 gases and the aerosol speciation for elemental and organic carbon (EC, OC), anhydrosugars, Cl(-), NO(3)(-), and 20 metals from 10 cooking fires, four garbage fires, three brick making kilns, three charcoal making kilns, and two crop residue fires. Global biofuel use has been estimated at over 2600 Tg/y. With several simple case studies we show that cooking fires can be a major, or the major, source of several gases and fine particles in developing countries. Insulated cook stoves with chimneys were earlier shown to reduce …

Laboratory Measurements Of Trace Gas Emissions From Biomass Burning Of Fuel Types From The Southeastern And Southwestern United States, I. R. Burling, Robert J. Yokelson, David W. T. Griffith, T. J. Johnson, P. Veres, J. M. Roberts, C. Warneke, S. P. Urbanski, J. Reardon, D. R. Weise, Wei Min Hao, J. De Gouw

Chemistry and Biochemistry Faculty Publications

Vegetation commonly managed by prescribed burning was collected from five southeastern and southwestern US military bases and burned under controlled conditions at the US Forest Service Fire Sciences Laboratory in Missoula, Montana. The smoke emissions were measured with a large suite of state-of-the-art instrumentation including an open-path Fourier transform infrared (OP-FTIR) spectrometer for measurement of gas-phase species. The OP-FTIR detected and quantified 19 gas-phase species in these fires: CO2, CO, CH4, C2H2, C2H4, C3H6, HCHO, HCOOH, CH3OH, CH3COOH, furan, H2O, NO, NO2, HONO, NH3, HCN, HCl, and SO2. Emission factors for these species are presented for each vegetation type burned. …

Evolution Of Trace Gases And Particles Emitted By A Chaparral Fire In California, S. K. Akagi, J. S. Craven, J. W. Taylor, G. R. Mcmeeking, Robert J. Yokelson, I. R. Burling, S. P. Urbanski, C. E. Wold, J. H. Seinfeld, H. Coe, M. J. Alvarado, D. R. Weise

Chemistry and Biochemistry Faculty Publications

Biomass burning (BB) is a major global source of trace gases and particles. Accurately representing the production and evolution of these emissions is an important goal for atmospheric chemical transport models. We measured a suite of gases and aerosols emitted from an 81 hectare prescribed fire in chaparral fuels on the central coast of California, US on 17 November 2009. We also measured physical and chemical changes that occurred in the isolated down-wind plume in the first similar to 4 h after emission. The measurements were carried out onboard a Twin Otter aircraft outfitted with an airborne Fourier transform infrared …

Nip/Duoxa Is Essential For Drosophila Embryonic Development And Regulates Oxidative Stress Response., Xiaojun Xie, Jack Hu, Xiping Liu, Hanjuan Qin, Anthony Percival-Smith, Yong Rao, Shawn S C Li

Biochemistry Publications

NIP/DuoxA, originally cloned as a protein capable of binding to the cell fate determinant Numb in Drosophila, was recently identified as a modulator of reactive oxygen species (ROS) production in mammalian systems. Despite biochemical and cellular studies that link NIP/DuoxA to the generation of ROS through the dual oxidase (Duox) enzyme, the in vivo function of NIP/DuoxA has not been characterized to date. Here we report a genetic and functional characterization of nip in Drosophila melanogaster. We show that nip is essential for Drosophila development as nip null mutants die at the 1(st) larval instar. Expression of UAS-nip, but not …

New Cyclophanes As Supramolecular Scaffolds: The Synthesis Of Tribenzo-1,4,7-Triazacyclononatriene., Andria M. Panagopoulos

Dissertations

Supramolecular chemistry involves the formation of complex molecular entities that have the capacity to participate in specific molecular recognition of guest molecules. A commonly employed scaffold in supramolecular chemistry is the trimeric crown-shaped molecule cyclotriveratrylene (CTV). CTV has been studied extensively for its capability of binding a number of smaller organic and organometallic guests within its bowl-shaped cleft and has been used as a building block enabling the construction of more complex cryptophanes. The goal of this research is the synthesis and characterization of a novel cyclophane, tribenzo-1,4,7-triazacyclononene and derivatives thereof. These new cyclophanes should have greater versatility than the …

Electroporation-Mediated Delivery Of A Naked Dna Plasmid Expressing Vegf To The Porcine Heart Enhances Protein Expression, W. G. Marshall Jr., B. A. Boone, J. D. Burgos, S. I. Gografe, M. K. Baldwin, M. L. Danielson, M. J. Larson, D. R. Caretto, Y. Cruz, B. Ferraro, L. C. Heller, K. E. Ugen, M. J. Jaroszeski, R. Heller

Bioelectrics Publications

Gene therapy is an attractive method for the treatment of cardiovascular disease. However, using current strategies, induction of gene expression at therapeutic levels is often inefficient. In this study, we show a novel electroporation (EP) method to enhance the delivery of a plasmid expressing an angiogenic growth factor (vascular endothelial growth factor, VEGF), which is a molecule previously documented to stimulate revascularization in coronary artery disease. DNA expression plasmids were delivered in vivo to the porcine heart with or without coadministered EP to determine the potential effect of electrically mediated delivery. The results showed that plasmid delivery through EP significantly …

Mzm1 Influences A Labile Pool Of Mitochondrial Zinc Important For Respiratory Function, Aaron Atkinson, Oleh Khalimonchuk, Pamela Smith, Hana Sabic, David Eide, Dennis R. Winge

Department of Biochemistry: Faculty Publications

Zinc is essential for function of mitochondria as a cofactor for

several matrix zinc metalloproteins. We demonstrate that a

labile cationic zinc component of low molecular mass exists in

the yeast mitochondrial matrix. This zinc pool is homeostatically

regulated in response to the cellular zinc status. This pool

of zinc is functionally important because matrix targeting of a

cytosolic zinc-binding protein reduces the level of labile zinc

and interferes with mitochondrial respiratory function. We

identified a series of proteins that modulate the matrix zinc

pool, one of which is a novel conserved mitochondrial protein

designated Mzm1. Mutant mzm1∆ cells …

Identification And Characterization Of Small Compound Inhibitors Of Human Fatp2, Angel Sandoval, Aalap Chokshi, Elliot D. Jesch, Paul N. Black, Concetta C. Dirusso

Department of Biochemistry: Faculty Publications

Fatty acid transport proteins (FATPs) are bifunctional proteins, which transport long chain fatty acids into cells and activate very long chain fatty acids by esterification with coenzyme A. In an effort to understand the linkage between cellular fatty acid transport and the pathology associated with excessive accumulation of exogenous fatty acids, we targeted FATP-mediated fatty acid transport in a high throughput screen of more than 100,000 small diverse chemical compounds in yeast expressing human FATP2 (hsFATP2). Compounds were selected for their ability to depress the transport of the fluorescent long chain fatty acid analogue, C₁-BODIPY-C₁₂. Among …

Evolution Of New Enzymatic Function By Structural Modulation Of Cysteine Reactivity In Pseudomonas Fluorescens Isocyanide Hydratase, Mahadevan Lakshminarasimhan, Peter Madzelan, Ruth Nan, Nicole Marie Milkovic, Mark A. Wilson

Department of Biochemistry: Faculty Publications

Isocyanide (formerly isonitrile) hydratase (EC 4.2.1.103) is an enzyme of the DJ-1 superfamily that hydrates isocyanides to yield the corresponding N-formamide. In order to understand the structural basis for isocyanide hydratase (ICH) catalysis, we determined the crystal structures of wild-type and several site-directed mutants of Pseudomonas fluorescens ICH at resolutions ranging from 1.0 to 1.9 Å. We also developed a simple UV-visible spectrophotometric assay for ICH activity using 2-naphthyl isocyanide as a substrate. ICH contains a highly conserved cysteine residue (Cys101) that is required for catalysis and interacts with Asp17, Thr102, and an ordered water molecule in the active …

The Roles Of Rhle And Hfq In Srna-Dependent Gene Regulation, Abeykoon Jayalath Iresha Sandeepanie Rathnayake

Wayne State University Theses

THE ROLES OF RHLE AND HFQ IN SRNA-DEPENDENT GENE REGULATION

by

ABEYKOON JAYALATH IRESHA SANDEEPANIE RATHNAYAKE

DECEMBER 2010

Advisor: Dr. Andrew Feig

Major: Chemistry

Degree: Master of Science

Bacteria are adapted to live in diverse environmental conditions. Thus, they show excellent tolerance and response to extreme environmental conditions caused by low or high temperatures, high salinity, reactive oxygen species or high nutrient concentrations. sRNAs have been identified and characterized as cis-acting or trans-acting post-transcriptional regulators in diverse cellular processes including virulence and adaptation to environmental stress (11,12,14,15). Interactions of sRNAs and target mRNAs result in translational repression, translational activation or/and …