Detection of Polyglutamine Expansion in a New Acidic Protein
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Polyq Disease: Misfiring of a Developmental Cell Death Program?
Review PolyQ disease: misfiring of a developmental cell death program? * * Elyse S. Blum , Andrew R. Schwendeman , and Shai Shaham Laboratory of Developmental Genetics, The Rockefeller University, 1230 York Avenue, New York, NY 10065, USA Polyglutamine (polyQ) repeat diseases are neurodegen- death [10]. The linker cell dies in the normal course of erative ailments elicited by glutamine-encoding CAG nu- C. elegans development, during gonadal morphogenesis of cleotide expansions within endogenous human genes. the male [11–13] (Figure 1). Linker cell death is indepen- Despite efforts to understand the basis of these diseases, dent of caspases and all other known apoptotic and necrotic the precise mechanism of cell death remains stubbornly C. elegans cell-death genes [13,14]. Mutations in the pqn- unclear. Much of the data seem to be consistent with a 41 gene block linker cell death, suggesting an important model in which toxicity is an inherent property of the role in linker cell demise. Furthermore, transcription of polyQ repeat, whereas host protein sequences surround- pqn-41 is induced immediately before the onset of cell ing the polyQ expansion modulate severity, age of onset, death, suggesting that this locus may be intimately con- and cell specificity. Recently, a gene, pqn-41, encoding a nected with the killing process [10]. pqn-41 encodes multi- glutamine-rich protein, was found to promote normally ple alternative transcripts, most of which can encode a occurring non-apoptotic cell death in Caenorhabditis ele- domain of 427 amino-acids, of which 35% are glutamines, gans. Here we review evidence for toxic and modulatory arranged in tracts of 1–8 residues in length [10]. -
Datasheet: VMA00301KT Product Details
Datasheet: VMA00301KT Description: TATA-BOX-BINDING PROTEIN ANTIBODY WITH CONTROL LYSATE Specificity: TATA-BOX-BINDING PROTEIN Format: Purified Product Type: PrecisionAb™ Monoclonal Isotype: IgG1 Quantity: 2 Westerns Product Details Applications This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visit www.bio-rad-antibodies.com/protocols. Yes No Not Determined Suggested Dilution Western Blotting 1/1000 PrecisionAb antibodies have been extensively validated for the western blot application. The antibody has been validated at the suggested dilution. Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Further optimization may be required dependant on sample type. Target Species Human Product Form Purified IgG - liquid Preparation 20μl Mouse monoclonal antibody prepared by affinity chromatography on Protein G from ascites Buffer Solution Phosphate buffered saline Preservative 0.09% Sodium Azide (NaN ) Stabilisers 3 Immunogen Purified His-tagged TATA-box-binding protein External Database Links UniProt: P20226 Related reagents Entrez Gene: 6908 TBP Related reagents Synonyms GTF2D1, TF2D, TFIID Specificity Mouse anti Human TATA-box-binding protein antibody recognizes the TATA-box-binding protein, -
Modifiers and Mechanisms of Multi-System Polyglutamine
c Indian Academy of Sciences REVIEW ARTICLE Modifiers and mechanisms of multi-system polyglutamine neurodegenerative disorders: lessons from fly models MOUSHAMI MALLIK and SUBHASH C. LAKHOTIA* Cytogenetics Laboratory, Department of Zoology, Banaras Hindu University, Varanasi 221 005, India Abstract Polyglutamine (polyQ) diseases, resulting from a dynamic expansion of glutamine repeats in a polypeptide, are a class of genetically inherited late onset neurodegenerative disorders which, despite expression of the mutated gene widely in brain and other tissues, affect defined subpopulations of neurons in a disease-specific manner. We briefly review the different polyQ- expansion-induced neurodegenerative disorders and the advantages of modelling them in Drosophila. Studies using the fly models have successfully identified a variety of genetic modifiers and have helped in understanding some of the molec- ular events that follow expression of the abnormal polyQ proteins. Expression of the mutant polyQ proteins causes, as a consequence of intra-cellular and inter-cellular networking, mis-regulation at multiple steps like transcriptional and post- transcriptional regulations, cell signalling, protein quality control systems (protein folding and degradation networks), axonal transport machinery etc., in the sensitive neurons, resulting ultimately in their death. The diversity of genetic modifiers of polyQ toxicity identified through extensive genetic screens in fly and other models clearly reflects a complex network effect of the presence of the mutated protein. Such network effects pose a major challenge for therapeutic applications. [Mallik M. and Lakhotia S. C. 2010 Modifiers and mechanisms of multi-system polyglutamine neurodegenerative disorders: lessons from fly models. J. Genet. 89, 497–526] Introduction of mutations which bring about expansion of unstable trinu- cleotide repeats in the genome. -
Control of Molecular Motor Motility in Electrical Devices
Control of Molecular Motor Motility in Electrical Devices Thesis submitted in accordance with the requirements of The University of Liverpool for the degree of Doctor in Philosophy By Laurence Charles Ramsey Department of Electrical Engineering & Electronics April 2014 i Abstract In the last decade there has been increased interest in the study of molecular motors. Motor proteins in particular have gained a large following due to their high efficiency of force generation and the ability to incorporate the motors into linear device designs. Much of the recent research centres on using these protein systems to transport cargo around the surface of a device. The studies carried out in this thesis aim to investigate the use of molecular motors in lab- on-a-chip devices. Two distinct motor protein systems are used to show the viability of utilising these nanoscale machines as a highly specific and controllable method of transporting molecules around the surface of a lab-on-a-chip device. Improved reaction kinetics and increased detection sensitivity are just two advantages that could be achieved if a motor protein system could be incorporated and appropriately controlled within a device such as an immunoassay or microarray technologies. The first study focuses on the motor protein system Kinesin. This highly processive motor is able to propel microtubules across a surface and has shown promise as an in vitro nanoscale transport system. A novel device design is presented where the motility of microtubules is controlled using the combination of a structured surface and a thermoresponsive polymer. Both topographic confinement of the motility and the creation of localised ‘gates’ are used to show a method for the control and guidance of microtubules. -
Molecular Biologist's Guide to Proteomics
Molecular Biologist's Guide to Proteomics Paul R. Graves and Timothy A. J. Haystead Microbiol. Mol. Biol. Rev. 2002, 66(1):39. DOI: 10.1128/MMBR.66.1.39-63.2002. Downloaded from Updated information and services can be found at: http://mmbr.asm.org/content/66/1/39 These include: REFERENCES This article cites 172 articles, 34 of which can be accessed free http://mmbr.asm.org/ at: http://mmbr.asm.org/content/66/1/39#ref-list-1 CONTENT ALERTS Receive: RSS Feeds, eTOCs, free email alerts (when new articles cite this article), more» on November 20, 2014 by UNIV OF KENTUCKY Information about commercial reprint orders: http://journals.asm.org/site/misc/reprints.xhtml To subscribe to to another ASM Journal go to: http://journals.asm.org/site/subscriptions/ MICROBIOLOGY AND MOLECULAR BIOLOGY REVIEWS, Mar. 2002, p. 39–63 Vol. 66, No. 1 1092-2172/02/$04.00ϩ0 DOI: 10.1128/MMBR.66.1.39–63.2002 Copyright © 2002, American Society for Microbiology. All Rights Reserved. Molecular Biologist’s Guide to Proteomics Paul R. Graves1 and Timothy A. J. Haystead1,2* Department of Pharmacology and Cancer Biology, Duke University,1 and Serenex Inc.,2 Durham, North Carolina 27710 INTRODUCTION .........................................................................................................................................................40 Definitions..................................................................................................................................................................40 Downloaded from Proteomics Origins ...................................................................................................................................................40 -
Protein Facility
Supporting core service facilities for biotechnology research by faculty, student, government, and industry scientists. More Office of Biotechnology at www.biotech.iastate.edu/service_facilities. Protein Facility Iowa State University’s Protein purification of proteins and of protein samples from 1D or 2D Facility provides expertise for the peptides can be accomplished. gels. Gel spots can be digested analysis, characterization, and with a variety of enzymes, and synthesis of proteins and peptides. Mass Spectrometry the resulting peptides can be After training, users can operate A matrix-assisted laser desorption/ analyzed to identify the protein. many instruments themselves. ionization time-of-flight (MALDI- TOF) mass spectrometer can be used SDS-PAGE / Electroblotting for determining the molecular weight The facility conducts sodium of proteins, peptides, glycoproteins, dodecyl sulfate polyacrylamide oligosaccharides, oligonucleotides, gel electrophoresis (SDS-PAGE) of and polymers. A quadrapole- proteins for purity and molecular TOF tandem mass spectrometer weight estimation. Gels can be is also available for obtaining electroblotted to nitrocellulose peptide sequence information. or to PVDF for immuno- detection and protein/peptide Peptide Synthesis sequencing, respectively. The facility can do both large- and small-scale peptide synthesis, 2-D Gel Electrophoresis including phosphopeptides, peptides The facility does two-dimensional with unusual amino acids, and electrophoresis by separating proteins multiple antigenic peptides (MAP). in the first dimension according to charge (isoelectric focusing), Circular Dichroism Protein and Peptide Sequencing followed by separating the focused Researchers who want to detect The Protein Facility provides proteins in the second dimension and quantitate the chirality of N-terminal protein/peptide sequence according to their molecular weight. -
Impact Factor: 3.958/ICV: 4.10 ISSN: 0978-7908 192 REVIEW ON: ELECTROPHORESIS: METHOD for PROTEIN SEPARATION Shindedipa
Impact factor: 3.958/ICV: 4.10 ISSN: 0978-7908 192 Pharma Science Monitor 7(2),Apr-Jun 2016 PHARMA SCIENCE MONITOR AN INTERNATIONAL JOURNAL OF PHARMACEUTICAL SCIENCES Journal home page: http://www.pharmasm.com REVIEW ON: ELECTROPHORESIS: METHOD FOR PROTEIN SEPARATION ShindeDipa V.*, JasminaSurati Department of Quality Assurance, Shree NaranjibhaiLalbhai Patel College of Pharmacy,Umrakh -394 345,Bardoli, Gujarat, India. ABSTRACT Electrophoresis is one of the widely used techniques in molecular biochemistry, microbiology, biomedical research. It is a type of protein separation method .It is one of the highly efficient techniques of analysis and sole method for separation of proteins for western blot, RNA studies etc. It is a both qualitative and quantitative analysis technique. Separation depend upon electrophoretic mobility.Electrophoresis technique are of various type like Moving boundary electrophoresis ,Zone electrophoresis ,Affinity electrophoresis ,Pulsed field electrophoresis ,Dielectrophoresis.this technique mainly used in antibiotic analysis,vaccine analysis DNA analysis and protein analysis as well as fingerprint analysis. KEYWORDS:Electrophoresis, Electrophoretic mobility,Zone Electrophoresis, Moving boundary Electrophoresis, Dielectricphoresis. INTRODUCTION Electrophoresis is a physical method of analysis based on the migration of electrically charged proteins, colloids, molecules or other particles dissolved or dispersed in an electrolyte solution in the direction of the electrode bearing the opposite polarity when an electric current is passed through it. Separations may be conducted in systems without support phases (such as free solution separation in capillary electrophoresis) or in stabilising media such as thin-later plates, filins or gels. The electrophoretic mobility is the rate of movement in metres per second of the charged particles under the action of an electric field of I volt per metre and is expressed in square metres per volt second. -
Bioanalytical Chemistry 8. Gel Electrophoresis and Blotting
91 Bioanalytical chemistry 8. Gel electrophoresis and blotting Suggested reading: Sections 9.1, 9.2.3, 9.2.4, 9.5.1, 10.1 to 10.7, 11.1 to 11.5, and 15.5 of Mikkelsen and Cortón, Bioanalytical Chemistry Primary Source Material • Chapter 4 and 6 of Biochemistry: Berg, Jeremy M.; Tymoczko, John L.; and Stryer, Lubert (NCBI bookshelf). • Chapter 3 and 7 of Molecular Cell Biology 4th ed. (Ch. 9, 5th ed.): Lodish, Harvey; Berk, Arnold; Zipursky, S. Lawrence; Matsudaira, Paul; Baltimore, David; Darnell, James E. (NCBI bookshelf). • Chapter 12 of Introduction to Genetic Analysis Anthony: J.F. Griffiths, Jeffrey H. Miller, David T. Suzuki, Richard C. Lewontin, William M. Gelbart (NCBI bookshelf). • Some animations are from http://www.wiley-vch.de/books/info/3-527-30300-6/. • Cancer examples from Weinberg, Robert (2007). The Biology of Cancer. Garland Science. • http://www.piercenet.com/ Electrophoresis 92 The velocity of migration (v) of a molecule in an electric field depends on the electric field strength (E), the net charge on the protein (z), and the frictional coefficient (f). Ez v = f The frictional coefficient f depends on both the mass and shape of the migrating molecule and the viscosity (η) of the medium. For a sphere of radius r, f = 6πηr € The speed of migration is therefore proportional to the charge:mass ratio. z Will the charge to mass ratio differ v ∝ between proteins? Between r different DNA molecules? € • Electrophoresis is a technique for separating, or resolving, molecules in a mixture under the influence of an applied electric field. -
Permease from Escherichia Coli
Plasticity of lipid-protein interactions in the function and topogenesis of the membrane protein lactose permease from Escherichia coli Mikhail Bogdanova,1, Philip Heacocka, Ziqiang Guanb, and William Dowhana,1 aDepartment of Biochemistry and Molecular Biology, University of Texas Medical School at Houston, Houston, TX, 77030; and bDepartment of Biochemistry, Duke University Medical Center, Durham, NC 27710 Edited by William T. Wickner, Dartmouth Medical School, Hanover, NH, and approved July 15, 2010 (received for review May 9, 2010) Phosphatidylcholine (PC) has been widely used in place of naturally N-terminal two-TM helical hairpin of PheP and GabP are in- occurring phosphatidylethanolamine (PE) in reconstitution of verted with respect to their orientation in PE-containing cells bacterial membrane proteins. However, PC does not support native and the remaining TMs. These permeases do not carry out pro- structure or function for several reconstituted transport proteins. ton-coupled energy-dependent uphill transport of substrate in Lactose permease (LacY) of Escherichia coli, when reconstituted cells lacking PE, but still display energy-independent downhill in E. coli phospholipids, exhibits energy-dependent uphill and transport. LacY reconstituted into total E. coli phospholipids car- energy-independent downhill transport function and proper con- ries out uphill transport with domains C6 and P7 on opposite formation of periplasmic domain P7, which is tightly linked to sides of the membrane bilayer as observed in wild-type cells uphill transport function. LacY expressed in cells lacking PE and (9). Leaving out PE during reconstitution results in only downhill containing only anionic phospholipids exhibits only downhill trans- transport with domains C6 and P7 residing on the same side of port and lacks native P7 conformation. -
Protein Blotting Guide
Electrophoresis and Blotting Protein Blotting Guide BEGIN Protein Blotting Guide Theory and Products Part 1 Theory and Products 5 Chapter 5 Detection and Imaging 29 Total Protein Detection 31 Transfer Buffer Formulations 58 5 Chapter 1 Overview of Protein Blotting Anionic Dyes 31 Towbin Buffer 58 Towbin Buffer with SDS 58 Transfer 6 Fluorescent Protein Stains 31 Stain-Free Technology 32 Bjerrum Schafer-Nielsen Buffer 58 Detection 6 Colloidal Gold 32 Bjerrum Schafer-Nielsen Buffer with SDS 58 CAPS Buffer 58 General Considerations and Workflow 6 Immunodetection 32 Dunn Carbonate Buffer 58 Immunodetection Workflow 33 0.7% Acetic Acid 58 Chapter 2 Methods and Instrumentation 9 Blocking 33 Protein Blotting Methods 10 Antibody Incubations 33 Detection Buffer Formulations 58 Electrophoretic Transfer 10 Washes 33 General Detection Buffers 58 Tank Blotting 10 Antibody Selection and Dilution 34 Total Protein Staining Buffers and Solutions 59 Semi-Dry Blotting 11 Primary Antibodies 34 Substrate Buffers and Solutions 60 Microfiltration (Dot Blotting) Species-Specific Secondary Antibodies 34 Stripping Buffer 60 Antibody-Specific Ligands 34 Blotting Systems and Power Supplies 12 Detection Methods 35 Tank Blotting Cells 12 Colorimetric Detection 36 Part 3 Troubleshooting 63 Mini Trans-Blot® Cell and Criterion™ Blotter 12 Premixed and Individual Colorimetric Substrates 38 Transfer 64 Trans-Blot® Cell 12 Immun-Blot® Assay Kits 38 Electrophoretic Transfer 64 Trans-Blot® Plus Cell 13 Immun-Blot Amplified AP Kit 38 Microfiltration 65 Semi-Dry Blotting Cells -
Wave Electroblotting, 2D, Complete Electrophoresis Systems And
Instruction Manual WAVE Standard and Tetrad Electroblotting, 2-D and Complete Electrophoresis Systems Electroblotting VS20WAVECBS AND WAVETETRAD1CBS Includes VS20WAVESYS and WAVEBI SW20 & WAVEBI 2-D Electrophoresis WAVEC2DS Includes VS20WAVESYS and VS20WAVEDCI VS20WAVEDC Complete (Electroblotting & 2-D Electrophoresis) VS20WAVECES AND WAVETETRAD1CES Includes VS20WAVECBS and VS20WAVEDCI 1 Contents:- Page 1) Safety Instructions 3 2) Packing Lists 4 3) Care and Maintenance 6 4) Usage Guidance and restrictions: 7 5) Set Up 8 6) Gel Casting 13 7) Gel Preparation 20 8) Gel Selection 20 9) Gel Pouring 22 10) Sample Preparation and Loading 23 11) Buffer Volume 25 12) Gel Running 25 13) Solutions 26 14) Vertical Gel Electrophoresis References 28 15) Combs 29 16) Protein Blotting using the WAVE 31 17) Buffer Volumes 32 18) Passive and Active Cooling 32 19) Run Conditions 34 20) Blotting References 35 21) Buffers 36 22) Blotting Troubleshooting 37 23) 1st Dimension Electrophoresis using the WAVEDCI Tube Gel Module 41 24) Capillary Tube Gel Pouring 41 25) 1st Dimension (IEF) Phase Tube Gel Running 43 26) 2-D, Size Determination Phase 44 27) Warranty 45 2 SAFETY PRECAUTION WHEN USED CORRECTLY, THESE UNITS POSE NO HEALTH RISK. HOWEVER, THESE UNITS CAN DELIVER DANGEROUS LEVELS OF ELECTRICITY AND ARE TO BE OPERATED ONLY BY QUALIFIED PERSONNEL FOLLOWING THE GUIDELINES LAID OUT IN THIS INSTRUCTION MANUAL. ANYONE INTENDING TO USE THIS EQUIPMENT SHOULD READ THE COMPLETE MANUAL THOROUGHLY. THE UNIT MUST NEVER BE USED WITHOUT THE SAFETY LID CORRECTLY IN POSITION. THE UNIT SHOULD NOT BE USED IF THERE IS ANY SIGN OF DAMAGE TO THE EXTERNAL TANK OR LID. -
Molecular Pathogenesis of Spinocerebellar Ataxia Type 6
Neurotherapeutics: The Journal of the American Society for Experimental NeuroTherapeutics Molecular Pathogenesis of Spinocerebellar Ataxia Type 6 Holly B. Kordasiewicz* and Christopher M. Gomez† *Ludwig Institute for Cancer Research, University of California at San Diego, La Jolla, California 92093; †Department of Neurology, University of Chicago, Chicago, Illinois 60637 Summary: Spinocerebellar ataxia type 6 (SCA6) is a neuro- to modulate or correct ion channel function. Alternatively, as a degenerative disorder caused by abnormal expansions of a disease in which the mutant protein contains an expanded poly- trinucleotide CAG repeat in exon 47 of the CACNA1A gene, glutamine tract, SCA6 may respond to the targets of drug which encodes the ␣1A subunit of the P/Q-type voltage-gated therapies developed for Huntington’s disease and other poly- calcium channel. The CAG repeat expansion is translated into glutamine disorders. In this review we will compare SCA6 to an elongated polyglutamine tract in the carboxyl terminus of other polyglutamine diseases and channelopathies, and we will ␣ ␣ the 1A subunit. The 1A subunit is the main pore-forming highlight recent advances in our understanding of ␣1A subunits subunit of the P/Q-type calcium channel. Patients with SCA6 and SCA6 pathology. We also propose a mechanism for how suffer from a severe form of progressive ataxia and cerebellar two seemingly divergent hypotheses can be combined into a dysfunction. Design of treatments for this disorder will depend cohesive model for disease progression. Key Words: Spino- on better definition of the mechanism of disease. As a disease cerebellar ataxia type 6 (SCA6), channelopathy, polyglutamine arising from a mutation in an ion channel gene, SCA6 may behave as an ion channelopathy, and may respond to attempts disease, P/Q-type calcium channel, Purkinje cell, cerebellum.