DOCSLIB.ORG

Methods in Biochemistry Metodologie Biochimiche Anno 2018-19 Stefania Brocca, Matilde Forcella, Paola Fusi

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Methods in Biochemistry Metodologie Biochimiche Anno 2018-19 Stefania Brocca, Matilde Forcella, Paola Fusi Methods in Biochemistry Metodologie Biochimiche Anno 2018-19 Stefania Brocca, Matilde Forcella, Paola Fusi [email protected] Tel. int.: 02 6448 3518 room: 5054, U3 - 5th floor https://elearning.unimib.it/ Metodologie Biochimiche 2018-19 01) Martedì 2/10 02) Venerdì 5/10 03) Martedì 9/10 Modulo frontale 04) Venerdì 12/10 Esercitazioni 4 crediti 05) Martedì 16/10 2 crediti (14 lezioni) 06) Venerdì 19/10 (20 h in 5 pomeriggi) 07) Martedì 23/10 Aule: U1-04; U1-10 08) Venerdì 26/10 3 turni da 1 settimana 09) Martedì 30/10 dicembre 2018, gennaio 2019, 10) Martedì 06/11 febbraio 2019 11) Venerdì 09/11 Lab 1026 12) Martedì 13/11 13) Venerdì 16/11 14) Martedì 20/11 Program (theoretical lessons) Protein «environments» Electrophoresis Hydrophobic effect Principles Solubility, salting in/salting out SDS-PAGE Hofmeister series (Blue)-native gel Denaturing agents Band-shift assay Dialysis Isoelectrophocusing Centrifugation techniques Bidimensional gel Principles Western blot Differential centrifugation Spectroscopies Density gradient centrifugation Principles Analytical ultracentrifugation Absorbance Chromatography Fluorescence Principles Circular dichroism Chromatographer set up Mass spectrometry Gel filtration Surface plasmon resonance Ion exchange Hydrophobic interactions Affinity tags Suggested references Wilson K. & Walker J. (2000) “Biochimica e Biologia Molecolare” Cortina, 2006 Numero di pagine: 777 Prezzo: ~ € 100 M. C. Bonaccorsi di Patti, R. Contestabile, M. L. Di Salvo Cited articles “Metodologie Biochimiche” Casa Editrice Ambrosiana, 2012 Numero di pagine: 289 Prezzo: ~ € 40 Protein preparation and analyses Cells/tissues Lysis Purification Characterization Protein extraction & purification Analitycal techniques (titration assays, structual analyses etc) Lessons 1-2 • Structure and stability of proteins • Protein denaturation • Hydrophobic effect • Ion solvatation • Protein solvatation • Hofmeister series • Protein precipitation-dialysis • Kinds of buffers Structure and stability of proteins Many factors can damage proteins • Temperature • Pressure • pH • Proteolytic activity • Reactive oxygen species (ROS) • Organic solvents • Urea/Guanidinium • Tannins Aim: keep protein native conditions Protein structure is stabilized by weak interactions • Hydrogen bonds • Van der Waals interactions • Electrostatic interactions • Hydrophobic effects It is not a direct attraction, but an «apparent force» that stems from entropy and involves solvent molecules Thermodynamics in protein folding ΔG = ΔH - TΔS Net difference of stability (∆G < 0) Hydrophobic effect (-T∆S << 0) In physiological conditions, ΔG <0. Low value! H bonds (∆H << 0) Conformational entropy Van der Waals of polypeptide chain interactions (∆H << 0) (-T∆S >>0) Proteins can be easily denatured!!! https://employees.csbsju.edu/hjakubowski/classes/ch331/protstructure/olhydrophobprot.html Hydrophobic or solvophobic effect • Hydrophobic effect: apolar compounds tend to aggregate in acqueus medium. Solvophobic effect applies to polar solvents different from water • The exclusion of non-polar groups from water is energetically favoured, (ΔG <0). Indeed, around non-polar groups, water molecules are more ordered (ΔS <0) water water Organic molecules One «hole» in water is better than two! ….. ….. Hydrophobic interactions and water structuring Native form Denatured form “quasifrozen” water patches about nonpolar molecules Frank and Evans, (1945) The journal of chemical physics. Hydrophobic interactions and water structuring Benzene «clathrates» Phe Hydrophobic effect lowers with temperature The transfer of a nonpolar solute into The transfer into hot water cold water (e.g. room temperature) is (e.g., boiling point) is opposed opposed by a large entropy. by a large enthalpy (needed to Dill et al., (2005). DOI:10.1146/annurev.biophys.34.040204.144517 boil water). Hydrophobic effect Summing up • Hydrophobic effect occurs when apolar compounds tend to aggregate in acqueus medium • Solvofobic effect is a hydrophobic effect occurring in a polar medium different from water • It is due to an «apparent» force (not direct attaction) • Entropic origin (exposition of apolar groups to water causes an ordered set-up among solvent molecules) Salt solutions Ionic strength (I ) 푛 1 Measure of charge 퐼 = 푐 푧 2 concentration in solution 2 푖 푖 푖=1 c = concentration, z = charge Es. NaCl 1 M I = 1/2 (1 . 12 + 1 . 12) = 2/2 = 1 . 2 . 2 Es. MgCl2 1 M I = 1/2 (1 2 + 2 1 ) = 6/2 = 3 • Salts affect both protein solubility and conformational stability • These effects depend on the kind of salt • Some salts can stabilize, others can denature proteins Water ordering Li+ and ionic radius - F Small Na+ + - Large Water K Cl Rb+ Br- Cs+ I- Water ordering depends on the ionic radius Bulk 2nd hydratation shell 1st hydratation shell Na+ Smaller the radius, thicker the shell Water ordering Li+ and ionic radius - F Small Na+ (kosmotropic ions) Large K+ Cl- Water (kaotropic ions) Rb+ Br- Cs+ I- Water ordering depends on the ionic radius Kosmotropic ions (ionic radius) Chaotropic ions For a given ionic radius, a negative ion interacts more strongly with water than a positive ion does The mystery of charge asimmetry - small ions - water water one H pointing outside F- Li+ More water dipoles and more both H pointing outside closer around the anion The mystery of charge asimmetry water - large ions - water Low charge density I- Cs+ • Weakly hydrated ions (bulk water–water bonds favored) • Tetrahedral water structures destabilized in their vicinity hydrophobic effect!! Larger ions are less hydrophilic Organization of water around a protein >ns ~ps protein surface inner How do salts affect protein solubility? Solubility of carboxyhemoglobin solubility in various electrolytes at 25 °C of Logarithm Salis e Ninham , DOI: 10.1039/C4CS00144C , Adattato da Green, J. Biol. Chem., 1932, 95 Root square of ion strength Low salt concentrations always favour solubility High salt concentrations can have different effects Poor solubility Salting in Salting out salt-solvent interactions Salting in Unfolding At low concentrationSalt concentration Protein-protein interactions salt-protein interactions At low concentration, salt screens out At concentr. > 0.1 M, salt competes for protein-protein charge interactions. water and removes the solvation sphere. Higher solubility Salting in “Salting out” or unfolding? Franz Hofmeister (1850-1922). First Faculty of Medicine, Charles University in Prague How do salts affect protein solubility? At salt concentration >0.1 M, certain kind of ions cause proteins precipitate out of solution (i.e., are salted out). Other types of salts cause proteins to become more soluble (salted in). Hofmeister series: ranking of ions for their ability to solubilize or precipitate proteins • Strong solvent interactions • Strong protein interactions • High interfacial tension • Lower interfacial tension • Native proteins precipitate due to • Proteins solubilized by interaction unavailability of solvent. Stabilizers with solvents. Denaturants Direct Hofmeister series Strong solvent Ion radius interactions ok Strong interactions Ion radius with protein… …depending on protein charges Electrostatic surface potential of a protein negative positive Direct Hofmeister series “Direct series”: empirically observed for proteins with mixed apolar and negatively charged surfaces “Indirect series”: reversed effects observed for proteins with mixed apolar and positively charged surfaces Isoelectric point (pI) = pH is the pH at which a molecule carries no net electrical charge in the statistical mean pI 10 pI 9 Protein A Protein B Electrostatic surface potentials of a protein pH 4.0 pH 7.0 pH 11.0 Several diverse Hofmeister series!! negative positive Electrostatic surface potential of three homologous proteinases K negative positive Helland et al. (2006) FEBS Journal 273:61–71 Schwierz et al. (2016), https://doi.org/10.1016/j.cocis.2016.04.003 Let’s start with some fixed points…. Guanidium chloride Tetramethylammonium Chaotropic ions = denaturing agents Guanidinium and other denaturants that lower solvophobic effect guanidinium (6 M) urea (8 M) Usually don’t cause Acetonitrile, methanol, ethanol, propanol etc aggregation Ammonium sulfate • Has a wide range of application • Proteins loose solubility as amm. sulfate concentration increases • Each protein precipitates at a particular concentration (empirically determined) Partial purification of all proteins with similar solubility characteristics. “Salt cuts”: fractionation of proteins from the same solution /extract Large-volume preparations Protein concentration (first steps of purification) Each protein precipitates at a particular ammonium sulfate concentration Protein response is specific and depends on composition, pI, size, quaternary structure etc. Scheme of «ammonium sulfate cut» 50% saturation ammonium sulfate (AS) pellet (contaminants) centrifugation supernatant 70% saturation AS pellet (target protein) centrifugation supernatant Ammonium sulfate precipitation table Grams for 1 L solution Final concentration (in % of solution saturation) concentration Starting Dialysis • Passage of solutes through a semi- permeable membrane. • Size of pores is defined in terms of cut-off (MW under which a solute can cross the membrane). • Protein stays in; water, salts, protein fragments, and other molecules smaller than the pore size pass through http://zoology.okstate.edu/zoo_lrc/biol1114/tutorials/Flash/Osmosis_Animation.htm Animation Protein minimal solubility is at pH = pI Aggregation at pH = pI Dialysis Dialysis is usually applied to buffer exchange. At the end of the procedure, the external buffer will substitute the internal one (internal components go outside and VICEVERSA). Video-procedure dialysis: http://www.youtube.com/watch?v=2Th0PuORsWY http://www.youtube.com/watch?v=y0ffJBFQswk http://www.youtube.com/watch?v=0Km2C2IwPLI .
Load more

Recommended publications
  • Specific Salt Effects on the Formation and Thermal Transitions Among Β-Lactoglobulin and Pectin Electrostatic Complexes
    Purdue University Purdue e-Pubs Open Access Theses Theses and Dissertations 2013 Specific altS Effects on the Formation and Thermal Transitions Among β-Lactoglobulin and Pectin Electrostatic Complexes Stacey Ann Hirt Purdue University, [email protected] Follow this and additional works at: https://docs.lib.purdue.edu/open_access_theses Part of the Agriculture Commons, Bioresource and Agricultural Engineering Commons, Food Science Commons, and the Physical Chemistry Commons Recommended Citation Hirt, Stacey Ann, "Specific altS Effects on the Formation and Thermal Transitions Among β-Lactoglobulin and Pectin Electrostatic Complexes" (2013). Open Access Theses. 34. https://docs.lib.purdue.edu/open_access_theses/34 This document has been made available through Purdue e-Pubs, a service of the Purdue University Libraries. Please contact [email protected] for additional information. Graduate School ETD Form 9 (Revised 12/07) PURDUE UNIVERSITY GRADUATE SCHOOL Thesis/Dissertation Acceptance This is to certify that the thesis/dissertation prepared Stacey A. Hirt By ! Entitled! Specific Salt Effects on the Formation and Thermal Transitions Among "-Lactoglobulin and Pectin Electrostatic Complexes! Master of Science For the degree of Is approved by the final examining committee: Owen Jones Chair Yuan Yao Ganesan Narsimhan To the best of my knowledge and as understood by the student in the Research Integrity and Copyright Disclaimer (Graduate School Form 20), this thesis/dissertation adheres to the provisions of Purdue University’s “Policy on Integrity
    [Show full text]
  • Protein Precipitation Using Ammonium Sulfate
    HHS Public Access Author manuscript Author ManuscriptAuthor Manuscript Author Curr Protoc Manuscript Author Protein Sci. Manuscript Author Author manuscript; available in PMC 2016 April 01. Published in final edited form as: Curr Protoc Protein Sci. 2001 May ; APPENDIX 3: Appendix–3F. doi:10.1002/0471140864.psa03fs13. Protein Precipitation Using Ammonium Sulfate Paul T. Wingfield Protein Expression Laboratory (HNB-27), National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS) - NIH, Bldg 6B, Room 1B130, 6 Center Drive, Bethesda MD 20814, Tel: 301-594-1313 Paul T. Wingfield: [email protected] Abstract The basic theory of protein precipitation by addition of ammonium sulfate is presented and the most common applications are listed, Tables are provided for calculating the appropriate amount of ammonium sulfate to add to a particular protein solution. Key terms for indexing Ammonium sulfate; ammonium sulfate tables; protein concentration; protein purification BASIC THEORY The solubility of globular proteins increases upon the addition of salt (<0.15 M), an effect termed salting-in. At higher salt concentrations, protein solubility usually decreases, leading to precipitation; this effect is termed salting-out ((Green and Hughes, 1955). Salts that reduce the solubility of proteins also tend to enhance the stability of the native conformation. In contrast, salting-in ions are usually denaturants. The mechanism of salting-out is based on preferential solvation due to exclusion of the cosolvent (salt) from the layer of water closely associated with the surface of the protein (hydration layer). The hydration layer, typically 0.3 to 0.4 g water per gram protein (Rupley et al., 1983), plays a critical role in maintaining solubility and the correctly folded native conformation.
    [Show full text]
  • Models and Mechanisms of Hofmeister Effects in Electrolyte
    Chem Soc Rev View Article Online REVIEW ARTICLE View Journal | View Issue Models and mechanisms of Hofmeister effects in electrolyte solutions, and colloid and protein Cite this: Chem. Soc. Rev., 2014, 43,7358 systems revisited Andrea Salis*ab and Barry W. Ninhamb Specific effects of electrolytes have posed a challenge since the 1880’s. The pioneering work was that of Franz Hofmeister who studied specific salt induced protein precipitation. These effects are the rule rather the exception and are ubiquitous in chemistry and biology. Conventional electrostatic theories (Debye– Hu¨ckel, DLVO, etc.) cannot explain such effects. Over the past decades it has been recognised that addi- tional quantum mechanical dispersion forces with associated hydration effects acting on ions are missing from theory. In parallel Collins has proposed a phenomenological set of rules (the law of matching water affinities, LMWA) which explain and bring to order the order of ion–ion and ion–surface site interactions at a qualitative level. The two approaches appear to conflict. Although the need for inclusion of quantum Creative Commons Attribution 3.0 Unported Licence. dispersion forces in one form or another is not questioned, the modelling has often been misleading and inappropriate. It does not properly describe the chemical nature (kosmotropic/chaotropic or hard/soft) of the interacting species. The success of the LMWA rules lies in the fact that they do. Here we point to the Received 30th April 2014 way that the two apparently opposing approaches might be reconciled. Notwithstanding, there are more DOI: 10.1039/c4cs00144c challenges, which deal with the effect of dissolved gas and its connection to ‘hydrophobic’ interactions, the problem of water at different temperatures and ‘water structure’ in the presence of solutes.
    [Show full text]
  • Are Hofmeister Series Relevant to Modern Ion-Specific Effects Research?
    Scholarly Research Exchange Volume 2008 • Article ID 818461 • doi:10.3814/2008/818461 Research Article Are Hofmeister Series Relevant to Modern Ion-Specific Effects Research? Terence J. Evens and Randall P. Niedz USDA, Agricultural Research Service, U.S. Horticultural Research Laboratory, 2001 South Rock Road, Ft. Pierce, FL 34945-3030, USA Correspondence should be addressed to Terence J. Evens, [email protected] Received 29 February 2008; Revised 17 April 2008; Accepted 5 May 2008 Ion-specific effects underlie a vast array of physicochemical and biological phenomena ranging from human physiology to biotechnology to ecology. These effects have traditionally been quantified by measuring the response of interest in a series of salt solutions at multiple concentrations; pH has consistently been shown to be of primary concern. However, salt-based approaches violate critical tenets of proper experimental design and introduce confounding errors that make it impossible to quantify ion- specific effects. For example, pH is a variable dependent on the type and concentration of ions in a solution, but is typically treated as an independent factor, thus confounding experiments designed to determine ion-specific effects. We examined the relevancy of ion-specific effects research in relation to these concepts and demonstrated how these ideas impact protein precipitation and enzyme activity. Based on these results, we present a conceptual and experimental framework of general applicability for proper quantification of ion-specific effects. Copyright © 2008 T. J. Evens and R. P. Niedz. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
    [Show full text]
  • Biochemistry 06 Salt Fractionation of Proteins
    . 1 Biochemical Techniques Biochemistry 06 Salt Fractionation of Proteins Description of Module Subject Name Biochemistry Paper Name 12 Biochemical Techniques Module Name/Title 06 Salt Fractionation of Proteins 2 Biochemical Techniques Biochemistry 06 Salt Fractionation of Proteins 1. Objectives Understanding the concept of protein fractionation Understanding protein fractionation with salt 2. Concept Map 3. Description 3.1 Protein purification Protein purification is vital for the characterization of protein structure, function and interactions, for example conformational alterations, substrate specificities, specific activities as well as interaction with other ligands. Further, for applications in the food and pharmaceutical industry, a high level of protein purity is 3 Biochemical Techniques Biochemistry 06 Salt Fractionation of Proteins essential and the desired protein must be purified over a number of steps. This is thus achieved through methods of protein purification. Purification is a multistep procedure. The various steps in the purification process are aimed removing non- protein components and impurities to finally separating out the desired protein in its pure form. 3.2 Fractionation of proteins The first step in purifying intracellular proteins is preparing a crude extract. The extract will contain a complex mixture of proteins from cell cytoplasm, and additional components such as macromolecules, cofactors and nutrients. The debris are removed by centrifugation and supernatant (crude protein extract) recovered. Crude preparations of extracellular proteins may be obtained by removing cells by centrifugation. The extract can be subjected to treatments that separate proteins into different fractions based on several properties such as size, charge etc. This process is known as fractionation. Fractionation helps in the removal of any other contaminating material and also in the enrichment of the desired protein fraction.
    [Show full text]
  • Open Dissertation 102714.Pdf
    The Pennsylvania State University The Graduate School Eberly College of Science ION SPECIFICTY AT THE POLYPEPTIDE BACKBONE: MOLECULAR MECHANISMS OF THE HOFMEISTER EFFECT A Dissertation in Chemistry by Kelvin Blaine Rembert II ©2014 Kelvin Blaine Rembert II Submitted in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy December 2014 i The dissertation of Kelvin B. Rembert II was reviewed and approved* by the following: Paul S. Cremer Professor of Chemistry & Biochemistry and Molecular Biology Dissertation Advisor Chair of Committee Scott A. Showalter Associate Professor of Chemistry William G. Noid Associate Professor of Chemistry Craig E. Cameron Professor of Biochemistry and Molecular Biology Barbara J. Garrison Shapiro Professor of Chemistry Head of Department of Chemistry *Signatures are on file in the graduate school ii ABSTRACT Ion Specificity at the Polypeptide Backbone: Molecular Level Mechanisms of the Hofmeister Effect (December 2014) Kelvin Blaine Rembert II, B.S., University of West Georgia, Carrollton, Georgia Chair of Advisory Committee: Dr. Paul S. Cremer The behavior of biomolecules in aqueous solutions is ion specific and follows the Hofmeister series. Discovered in 1888, the Hofmeister series is a recurring trend that originally ranked cations and anions by their ability to precipitate proteins from aqueous salt solutions. Over the course of the 21st century however, the Hofmeister effect was found to extend well beyond protein precipitation studies and occurs in myriad bio-chemical and physico-chemical systems. Today, the field remains an active area of research and has gained the attention of many researchers due to the elusiveness in developing a theoretical framework for predicting ion specific effects in water.
    [Show full text]
  • Collaborative Routes to Clarifying the Murky Waters of Aqueous Supramolecular Chemistry Paul S
    REVIEW ARTICLE PUBLISHED ONLINE: 19 DECEMBER 2017 | DOI: 10.1038/NCHEM.2894 Collaborative routes to clarifying the murky waters of aqueous supramolecular chemistry Paul S. Cremer1*, Amar H. Flood2*, Bruce C. Gibb3* and David L. Mobley4* On planet Earth, water is everywhere: the majority of the surface is covered with it; it is a key component of all life; its vapour and droplets fill the lower atmosphere; and even rocks contain it and undergo geomorphological changes because of it. A community of physical scientists largely drives studies of the chemistry of water and aqueous solutions, with expertise in biochemistry, spectroscopy and computer modelling. More recently, however, supramolecular chemists — with their expertise in macrocyclic synthesis and measuring supramolecular interactions — have renewed their interest in water-mediated non- covalent interactions. These two groups offer complementary expertise that, if harnessed, offer to accelerate our understanding of aqueous supramolecular chemistry and water writ large. This Review summarizes the state-of-the-art of the two fields, and highlights where there is latent chemical space for collaborative exploration by the two groups. ater is ubiquitous and essential to life on planet Earth. A and a good HB acceptor (β = 0.47). Through the combination of its full understanding of aqueous solutions is therefore of permanent dipole, electron-deficient hydrogens and lone pairs, water Wsignificance to a wide range of fields within the atmos- forms strong electrostatic interactions with other waters and ions, pheric, environmental, biological and geological sciences. Within but weaker interactions with less polar solutes. the chemical sciences, a deeper appreciation of how non-covalent In the crystalline solid state, each water molecule makes four interactions and chemical transformations are influenced by water HBs with its neighbours; however, in the liquid state near room would benefit a variety of fields.
    [Show full text]
  • Interactions Between Hofmeister Anions and the Binding Pocket of a Protein † † ∥ ⊥ ¶ Jerome M
    Article pubs.acs.org/JACS Interactions between Hofmeister Anions and the Binding Pocket of a Protein † † ∥ ⊥ ¶ Jerome M. Fox, Kyungtae Kang, Woody Sherman, Annie Heroux,́ G. Madhavi Sastry, † † ▽ † ‡ § Mostafa Baghbanzadeh, Matthew R. Lockett, , and George M. Whitesides*, , , † Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, Massachusetts 02138, United States ‡ Wyss Institute for Biologically Inspired Engineering, Harvard University, 60 Oxford Street, Cambridge, Massachusetts 02138, United States § The Kavli Institute for Bionano Science and Technology, Harvard University, 29 Oxford Street, Cambridge, Massachusetts 02138, United States ∥ Schrödinger, 120 West 45th Street, New York, New York 10036, United States ⊥ Photon Science Division, Energy Sciences Directorate, Brookhaven National Laboratory, Building 745, Upton, New York 11937, United States ¶ Schrödinger, Sanali Infopark, 8-2-120/113 Banjara Hills, Hyderabad 11937, Andhra Pradesh, India *S Supporting Information ABSTRACT: This paper uses the binding pocket of human carbonic anhydrase II (HCAII, EC 4.2.1.1) as a tool to examine the properties of Hofmeister anions that determine (i) where, and how strongly, they associate with concavities on the surfaces of proteins and (ii) how, upon binding, they alter the structure of water within those concavities. Results from X-ray crystallography and isothermal titration calorimetry show that most anions associate with the binding pocket of HCAII by forming inner-sphere ion pairs with the Zn2+ cofactor. In these ion pairs, the free energy of anion−Zn2+ association is inversely proportional to the free energetic cost of anion dehydration; this relationship is consistent with the mechanism of ion pair formation suggested by the “law of matching water affinities”.
    [Show full text]
  • 1 Water Runs Deep
    1 1 Water Runs Deep Nicholas E. Ernst and Bruce C. Gibb Tulane University, Department of Chemistry, 2015 Percival Stern Hall, 6400 Freret Street, New Orleans, LA, 70118, USA 1.1 ­The Control of Water There is current consensus among archeology scholars that there are five cradles of civilization, geographical locations where the first civilizations emerged thousands of years ago. These five areas – the Fertile Crescent, the Indus River, the Yellow River, the Central Andes, and Mesoamerica – each independently gave rise to a new level of human existence [1–3]. What do these sites have in common? They all shared a combination of favorable geography and the opportunity to control water. Because of its central role in life on Earth, the idea of controlling water mani- fests in many aspects of human endeavor. In agriculture, the key is irrigation – how to bring water to the fertile land and crops. “Controlling” the oceans and rivers requires harbors, moorings, and boats to open up trading routes or the possibility of fishing, while controlling sewage has been key to the success of cities. Controlling water also allowed energy production in the form of water- mills and, in more modern times, dams, tidal barrages, and wave energy con- vertors. Unfortunately, the control of water can also be used to justify war. Conflicts between nations, states, or groups, for example, the violence in the war in Sudanese Darfur, can be partially attributed to water [4, 5]. For many different reasons, the control of water at the molecular and nanoscale levels is also key. Many scientific disciplines work toward being able to under- stand and control water purification for waste treatment and desalination [6, 7], water flow for microfluidics and ultra‐hydrophobic materials [8], and solvation to in turn control dissolved matter.
    [Show full text]
  • The Effect of Hofmeister Series Counterions on the Colloidal and Antimicrobial Properties of Triple- Headed Cationic Amphiphiles Kirstie A
    James Madison University JMU Scholarly Commons Senior Honors Projects, 2010-current Honors College Spring 2016 The effect of Hofmeister series counterions on the colloidal and antimicrobial properties of triple- headed Cationic amphiphiles Kirstie A. Thompson James Madison University Follow this and additional works at: https://commons.lib.jmu.edu/honors201019 Part of the Organic Chemistry Commons Recommended Citation Thompson, Kirstie A., "The effect of Hofmeister series counterions on the colloidal and antimicrobial properties of triple-headed Cationic amphiphiles" (2016). Senior Honors Projects, 2010-current. 198. https://commons.lib.jmu.edu/honors201019/198 This Thesis is brought to you for free and open access by the Honors College at JMU Scholarly Commons. It has been accepted for inclusion in Senior Honors Projects, 2010-current by an authorized administrator of JMU Scholarly Commons. For more information, please contact [email protected]. The Effect of Hofmeister Series Counterions on the Colloidal and Antimicrobial Properties of Triple-Headed Cationic Amphiphiles _______________________ An Honors Program Project Presented to the Faculty of the Undergraduate College of Science and Mathematics James Madison University _______________________ by Kirstie Ann Thompson May 2016 Accepted by the faculty of the Department of Chemistry and Biochemistry, James Madison University, in partial fulfillment of the requirements for the Honors Program. FACULTY COMMITTEE: HONORS PROGRAM APPROVAL: Project Advisor: Kevin L. Caran, Ph.D. Bradley R. Newcomer, Ph.D., Associate Professor, Chemistry Director, Honors Program Reader: Yanjie Zhang, Ph.D. Assistant Professor, Chemistry Reader: Kyle Seifert, Ph.D. Associate Professor, Biology PUBLIC PRESENTATION This work is accepted for presentation, in part or in full, at James Madison University Department of Chemistry and Biochemistry Spring Symposium on April 14th 2016 .
    [Show full text]
  • Hydration of Cations: a Key to Understanding of Specific Cation Effects on Aggregation Behaviors of PEO-PPO-PEO Triblock Copolymers Jacob C
    Article pubs.acs.org/JPCB Hydration of Cations: A Key to Understanding of Specific Cation Effects on Aggregation Behaviors of PEO-PPO-PEO Triblock Copolymers Jacob C. Lutter, Tsung-yu Wu, and Yanjie Zhang* Department of Chemistry and Biochemistry, James Madison University, Harrisonburg, Virginia 22807, United States ABSTRACT: This work reports results from the interactions of a series of monovalent and divalent cations with a triblock copolymer, poly(ethylene oxide)-b-poly(propylene oxide)-b- poly(ethylene oxide) (PEO-PPO-PEO). Phase transition temperatures of the polymer in the presence of chloride salts with six monovalent and eight divalent cations were measured using an automated melting point apparatus. The polymer undergoes a two-step phase transition, consisting of micellization of the polymer followed by aggregation of the micelles, in the presence of all the salts studied herein. The results suggest that hydration of cations plays a key role in determining the interactions between the cations and the polymer. The modulation of the phase transition temperature of the polymer by cations can be explained as a balance between three interactions: direct binding of cations to the oxygen in the polymer chains, cations sharing one water molecule with the polymer in their hydration layer, and cations interacting with the polymer via two water molecules. + + + + + + Monovalent cations Na ,K,Rb, and Cs do not bind to the polymer, while Li and NH4 and all the divalent cations investigated including Mg2+,Ca2+,Sr2+,Ba2+,Co2+,Ni2+,Cu2+, and Cd2+ bind to the polymer. The effects of the cations correlate well with their hydration thermodynamic properties.
    [Show full text]
  • Protein Precipitation for the Purification of Therapeutic Proteins
    Protein Precipitation for the Purification of Therapeutic Proteins by Christopher W Morris A thesis submitted for the degree of Doctor of Engineering in The Department of Biochemical Engineering, UCL 1 Abstract This thesis documents the application of precipitant scouting and analytical tools for the development of a precipitation process for the purification of therapeutic proteins in biopharmaceuticals. Precipitation has the potential to bypass the bottlenecks in productivity experienced with packed bed chromatographic separations, offering a fast, robust first purification step with volume reduction at low cost. In order to evaluate a large number of precipitation candidates, microscale investigations aided by automated liquid handling robotics were chosen, which conferred precision, speed, and complex experimental design with microliter material requirements and in-line analytics. A precipitation screening methodology was successfully built onto a Tecan liquid handling platform including product recovery and techniques for measuring soluble protein, liquid volumes, and recovered protein precipitate. The protocol was supplemented with a custom- build Excel VBA driven Tecan control tool. This accelerated progress by freeing up the potential of the liquid handling arm, which was curtailed by system software. These techniques were then used to characterise effective precipitants for the purification of monoclonal antibodies. Optimal conditions were identified on pure protein models, which were the bridged to process relevant cell culture fluid. Process performance, notably recovery yield and product quality were the investigated on the precipitant conditions brought forward. Process integration aspects were explored, by linking precipitation with an anion exchange chromatography step. This led to discussion on future work and process scale up considerations. 2 Impact Statement This impact statement serves as a suitable place to reflect on the work presented in this thesis and how it relates to academic and industrial applications.
    [Show full text]