DOCSLIB.ORG

Evaluating Protein Structure and Dynamics Using Co-Solvents, Photochemical Triggers, and Site-Specific Spectroscopic Probes

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Evaluating Protein Structure and Dynamics Using Co-Solvents, Photochemical Triggers, and Site-Specific Spectroscopic Probes University of Pennsylvania ScholarlyCommons Publicly Accessible Penn Dissertations 2017 Evaluating Protein Structure And Dynamics Using Co-Solvents, Photochemical Triggers, And Site-Specific Spectroscopic Probes Rachel Marie Abaskharon University of Pennsylvania, [email protected] Follow this and additional works at: https://repository.upenn.edu/edissertations Part of the Physical Chemistry Commons Recommended Citation Abaskharon, Rachel Marie, "Evaluating Protein Structure And Dynamics Using Co-Solvents, Photochemical Triggers, And Site-Specific Spectroscopic Probes" (2017). Publicly Accessible Penn Dissertations. 2154. https://repository.upenn.edu/edissertations/2154 This paper is posted at ScholarlyCommons. https://repository.upenn.edu/edissertations/2154 For more information, please contact [email protected]. Evaluating Protein Structure And Dynamics Using Co-Solvents, Photochemical Triggers, And Site-Specific Spectroscopic Probes Abstract As ubiquitous and diverse biopolymers, proteins are dynamic molecules that are constantly engaging in inter- and intramolecular interactions responsible for their structure, fold, and function. Because of this, gaining a comprehensive understanding of the factors that control protein conformation and dynamics remains elusive as current experimental techniques often lack the ability to initiate and probe a specific interaction or conformational transition. For this reason, this thesis aims to develop methods to control and monitor protein conformations, conformational transitions, and dynamics in a site-specific manner, as well as to understand how specific and non-specific interactions affect the protein folding energy landscape. First, by using the co-solvent, trifluoroethanol (TFE), we show that the rate at which a peptide folds can be greatly impacted and thus controlled by the excluded volume effect. Secondly, we demonstrate the utility of several light-responsive molecules and reactions as methods to manipulate and investigate protein-folding processes. Using an azobenzene linker as a photo-initiator, we are able to increase the folding rate of a protein system by an order of magnitude by channeling a sub-population through a parallel, faster folding pathway. Additionally, we utilize a tryptophan-mediated electron transfer process to a nearby disulfide bond to strategically unfold a protein molecule with ultraviolet light. We also demonstrate the potential of two ruthenium polypyridyl complexes as ultrafast phototriggers of protein reactions. Finally, we develop several site-specific spectroscopic probes of protein structure and environment. Specifically, we demonstrate that a 13C-labeled aspartic acid residue constitutes a useful site-specific infrared probe for investigating salt-bridges and hydration dynamics of proteins, particularly in proteins containing several acidic amino acids. We also show that a proline-derivative, 4-oxoproline, possesses novel infrared properties that can be exploited to monitor the cis-trans isomerization process of individual proline residues in proteins. Degree Type Dissertation Degree Name Doctor of Philosophy (PhD) Graduate Group Chemistry First Advisor Feng Gai Keywords Co-Solvents, Infrared Spectroscopy, Photochemical Triggers, Protein Folding, Site-Specific Spectroscopic Probes Subject Categories Physical Chemistry This dissertation is available at ScholarlyCommons: https://repository.upenn.edu/edissertations/2154 EVALUATING PROTEIN STRUCTURE AND DYNAMICS USING CO-SOLVENTS, PHOTOCHEMICAL TRIGGERS, AND SITE-SPECIFIC SPECTROSCOPIC PROBES Rachel M. Abaskharon A DISSERTATION in Chemistry Presented to the Faculties of the University of Pennsylvania in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy 2017 Supervisor of Dissertation ________________________ Feng Gai Edmund J. and Louise W. Kahn Endowed Term Professor of Chemistry Graduate Group Chairperson ________________________ Gary A. Molander Hirschmann-Makineni Professor of Chemistry Dissertation Committee Marsha I. Lester (chair), Edmund J. Kahn Distinguished Professor Tobias Baumgart, Associate Professor of Chemistry, Graduate Chair Sergei Vinogradov, Associate Professor of Biochemistry and Biophysics EVALUATING PROTEIN STRUCTURE AND DYNAMICS USING CO-SOLVENTS, PHOTOCHEMICAL TRIGGERS, AND SITE-SPECIFIC SPECTROSCOPIC PROBES COPYRIGHT 2017 Rachel M. Abaskharon This work is licensed under the Creative Commons Attribution- NonCommercial-ShareAlike 3.0 License To view a copy of this license, visit https://creativecommons.org/licenses/by-nc-sa/3.0/us/ DEDICATION To my Nanny: You always believed in me even when I did not believe in myself iii ACKNOWLEDGEMENTS The past five years at Penn have proved to be challenging and rewarding, educational and inspiring, confidence building and fulfilling. I came into graduate school as an excited yet naïve chemist, ready to see where my research would take me. I am leaving, still excited, but much wiser, knowing that I now possess the skills, knowledge, and experience to follow my dreams to my full capability. Professor Feng Gai has been instrumental in helping me develop into the chemist I am today. He took a chance on an eager, new college graduate and saw the potential in me, early on, urging me to stretch myself academically and professionally, while showing his support in my endeavors. His trust in my abilities provided me with the confidence I needed to tackle any obstacle and push myself beyond the limits I thought I had. I am very grateful for his time, dedication, insight, and guidance throughout my time in his lab, which has been invaluable in allowing me to accomplish what I have today. I would also like to thank my committee members, collaborators and labmates for all of their guidance and support. Professors Marsha Lester, Tobias Baumgart, and Sergei Vinogradov have not only showed interest in my research but have also encouraged me in my scientific accomplishments and goals. I value the discussions I have had with each of them which have allowed to expand my own perspective of my work. Additionally, I appreciate the efforts of Dr. Jeffery Saven in helping me secure external funding throughout my time at Penn. My collaborators and labmates deserve a great amount in credit all of the work here. The collaborations of Professor G. Andrew Woolley, Dr. Stephen Brown, Professor Amos B. Smith III, Teresa Rapp, and Professor Ivan Dmochowski have helped tremendously with the synthesis of compounds and proteins, iv protein bioconjugation, and scientific insights for which I am very grateful. Additionally, I would like to thank all the former and current Gai lab group members and the UOPL staff members that I have had the pleasure of working with including: Dr. Robert Culik, Dr. Ileana Pazos, Dr. Beatrice Markiewicz, Dr. Chun-Wei Lin, Dr. Wenkai Zhang, Dr. Bei Ding, Dr. Kwang-Im Oh, Dr. Natalia Rubtsova, Dr. Patrick Walsh, Dr. Thomas Troxler, Dr. Jian-Xin Chen, Mary Rose Hilaire, Ismail Ahmed, Jeffrey Rodgers, Debopreeti Mukherjee, and Arusha Acharyya. I have been blessed with working with such a talented, kind and inspirational group of people, and I am sincerely thankful for all of your help and friendships. In particular, I want to thank Drs. Robert Culik, Ileana Pazos, and Beatrice Markiewicz for training, mentoring and guiding me in lab and life. Finally, I would never have accomplished what I have today if not for my friends and family supporting me. I am truly thankful to my friends, Melanie Hutnick and Leigh Matano, who were always there through the ups and downs of grad school despite the distance that separated us. To my sister-from-a-different-mister, Dr. Michelle Bunagan, I would not be here without you sharing your love of thermodynamics with me and encouraging me to pursue my Ph.D. I am very thankful to my parents, William and Carol Roesch, who taught me at a young age that in all I do, I should do it with all of my heart and ability to the glory of God. I also want to thank my “little” sister, Jillian Roesch, for making sure I grew up okay. To my husband, Ray Abaskharon, thank you for helping me chase my dreams and believing that “it’s you and me against the world, and the world has nothing on us.” Lastly and most importantly, I want to thank God for guiding me throughout my whole time here and blessing me with everything I have. v ABSTRACT EVALUATING PROTEIN STRUCTURE AND DYNAMICS USING CO-SOLVENTS, PHOTOCHEMICAL TRIGGERS, AND SITE-SPECIFIC SPECTROSCOPIC PROBES Rachel M. Abaskharon Feng Gai As ubiquitous and diverse biopolymers, proteins are dynamic molecules that are constantly engaging in inter- and intramolecular interactions responsible for their structure, fold, and function. Because of this, gaining a comprehensive understanding of the factors that control protein conformation and dynamics remains elusive as current experimental techniques often lack the ability to initiate and probe a specific interaction or conformational transition. For this reason, this thesis aims to develop methods to control and monitor protein conformations, conformational transitions, and dynamics in a site-specific manner, as well as to understand how specific and non-specific interactions affect the protein folding energy landscape. First, by using the co-solvent, trifluoroethanol (TFE), we show that the rate at which a peptide folds can be greatly impacted and thus controlled by the excluded volume effect. Secondly, we demonstrate the utility of several light-responsive molecules and reactions as methods to manipulate
Load more

Recommended publications
  • Determination of Pk, Values of the Histidine Side
    Protein Science (1997), 6:1937-1944. Cambridge University Press. Printed in the USA Copyright 0 1997 The Protein Society Determination of pK, values of the histidine side chains of phosphatidylinositol-specific phospholipase C from Bacillus cereus by NMR spectroscopy and site-directed mutagenesis TUN LIU, MARGRET RYAN, FREDERICK W. DAHLQUIST, AND 0. HAYES GRIFFITH Institute of Molecular Biology and Department of Chemistry, University of Oregon, Eugene, Oregon 97403 (RECEIVEDDecember 4, 1996: ACCEPTEDMay 19, 1997) Abstract Two active site histidine residues have been implicated in the catalysis of phosphatidylinositol-specific phospholipase C (PI-PLC). In this report, we present the first study of the pK,, values of histidines of a PI-PLC. All six histidines of Bacillus cereus PI-PLC were studied by 2D NMR spectroscopy and site-directed mutagenesis. The protein was selec- tively labeled with '3C"-histidine. A series of 'H-I3C HSQC NMR spectra were acquired over a pH range of 4.0-9.0. Five of the six histidines have been individually substituted with alanine to aid the resonance assignments in the NMR spectra. Overall, the remaining histidines in the mutants show little chemical shift changes in the 'H-"C HSQC spectra, indicating that the alanine substitution has no effect on the tertiary structure of the protein. H32A and H82A mutants are inactive enzymes, while H92A and H61A are fully active, and H81A retains about 15% of the wild-type activity. The active site histidines, His32 and His82, display pK,, values of 7.6 and 6.9, respectively. His92 and His227 exhibit pK, values of 5.4 and 6.9.
    [Show full text]
  • Families and the Structural Relatedness Among Globular Proteins
    Protein Science (1993), 2, 884-899. Cambridge University Press. Printed in the USA. Copyright 0 1993 The Protein Society -~~ ~~~~ ~ Families and the structural relatedness among globular proteins DAVID P. YEE AND KEN A. DILL Department of Pharmaceutical Chemistry, University of California, San Francisco, California94143-1204 (RECEIVEDJanuary 6, 1993; REVISEDMANUSCRIPT RECEIVED February 18, 1993) Abstract Protein structures come in families. Are families “closely knit” or “loosely knit” entities? We describe a mea- sure of relatedness among polymer conformations. Based on weighted distance maps, this measure differs from existing measures mainly in two respects: (1) it is computationally fast, and (2) it can compare any two proteins, regardless of their relative chain lengths or degree of similarity. It does not require finding relative alignments. The measure is used here to determine the dissimilarities between all 12,403 possible pairs of 158 diverse protein structures from the Brookhaven Protein Data Bank (PDB). Combined with minimal spanning trees and hier- archical clustering methods,this measure is used to define structural families. It is also useful for rapidly searching a dataset of protein structures for specific substructural motifs.By using an analogy to distributions of Euclid- ean distances, we find that protein families are not tightly knit entities. Keywords: protein family; relatedness; structural comparison; substructure searches Pioneering work over the past 20 years has shown that positions after superposition. RMS is a useful distance proteins fall into families of related structures (Levitt & metric for comparingstructures that arenearly identical: Chothia, 1976; Richardson, 1981; Richardson & Richard- for example, when refining or comparing structures ob- son, 1989; Chothia & Finkelstein, 1990).
    [Show full text]
  • Genomics of an Extreme Psychrophile, Psychromonas
    BMC Genomics BioMed Central Research article Open Access Genomics of an extreme psychrophile, Psychromonas ingrahamii Monica Riley*1, James T Staley2, Antoine Danchin3, Ting Zhang Wang3, Thomas S Brettin4, Loren J Hauser5, Miriam L Land5 and Linda S Thompson4 Address: 1Bay Paul Center, Marine Biological Laboratory, Woods Hole, MA 02543, USA, 2University of Washington, Seattle, WA 98195-7242, USA, 3Genetics of Bacterial Genomes, CNRS URA2171, Institut Pasteur, 28 rue du Dr Roux, 75015 Paris, France, 4DOE Joint Genome Institute, Bioscience Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USA and 5Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA Email: Monica Riley* - [email protected]; James T Staley - [email protected]; Antoine Danchin - [email protected]; Ting Zhang Wang - [email protected]; Thomas S Brettin - [email protected]; Loren J Hauser - [email protected]; Miriam L Land - [email protected]; Linda S Thompson - [email protected] * Corresponding author Published: 6 May 2008 Received: 3 September 2007 Accepted: 6 May 2008 BMC Genomics 2008, 9:210 doi:10.1186/1471-2164-9-210 This article is available from: http://www.biomedcentral.com/1471-2164/9/210 © 2008 Riley et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background: The genome sequence of the sea-ice bacterium Psychromonas ingrahamii 37, which grows exponentially at -12C, may reveal features that help to explain how this extreme psychrophile is able to grow at such low temperatures.
    [Show full text]
  • United States Patent (19) 11) 4,039,382 Thang Et Al
    United States Patent (19) 11) 4,039,382 Thang et al. 45 Aug. 2, 1977 54 MMOBILIZED RIBONUCLEASE AND -Enzyme Systems, Journal of Food Science, vol. 39, ALKALINE PHOSPHATASE 1974, (pp. 647-652). 75 Inventors: Minh-Nguy Thang, Bagneux; Annie Zaborsky, O., Immobilized Enzymes, CRC Press, Guissani born Trachtenberg, Fresnes, Cleveland, Ohio, 1973, (pp. 124-126). both of France Primary Examiner-David M. Naff 73 Assignee: Choay S. A., Paris, France Attorney, Agent, or Firm-Browdy and Neimark 21 Appl. No.: 678,459 22 Filed: Apr. 19, 1976 57 ABSTRACT An insoluble, solid matrix carrying simultaneously sev 30 Foreign Application Priority Data eral different enzymatic functions, is constituted by the Apr. 23, 1975 France ................................ 75.12667 conjoint association by irreversible binding on a previ ously activated matrix support, of a nuclease selected 51) int. Cl? ........................... C07G 7/02; C12B 1/00 from the group of ribonucleases A, T, T, U, and an 52 U.S. Cl. ................................... 195/28 N; 195/63; alkaline phosphatease. Free activated groups of the 195/68; 195/DIG. 11; 195/116 matrix after binding of the enzymes, are neutralized by 58 Field of Search ................... 195/63, 68, DIG. 11, a free amino organic base. The support is selected from 195/116, 28 N among non-denaturing supports effecting the irrevers 56) References Cited ible physical adsorption of the enzymes, such as sup ports of glass or quartz beads, highly cross-linked gels PUBLICATIONS of the agarose or cellulose type. Polymers AUott, A. Lee, J. C., Preparation and Properties of Water Insolu Cott, AGot and/or oligonucleotides U, C, A or G, of ble Derivatives of Ribonuclease Ti.
    [Show full text]
  • Apr 1 0 1981 Ubraries 2
    Mechanism of DNA Chain Initiation by the dnaG Protein of Escherichia coli by Daniel Jeffrey Capon B.S., Massachusetts Institute of Technology (1976) SUBMITTED IN PARTIAL FULFILL1ENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY at the MASSACHUSETTS INSTITUTE OF TECHNOLOGY January 1981 0) Massachusetts Institute of Technology 1981 Signature of Author__ %A~A(Departjent of Biology Certified by A U Thesis Supervisor Accepted by ~'RCHIVES Chairman, Departmental Committee MASSACHUSM INSTiTUTE OF TECHNOL1Y APR 1 0 1981 UBRARIES 2 Mechanism of DNA Chain Initiation by the dnaG protein of Escherichia coli by Daniel Jeffrey Capon Submitted to the Department of Biology on January 28, 1981 in partial fulfillment of the requirements for the degree of Doctor of Philosophy ABSTRACT All known DNA polymerases are unable to initiate the synthesis of DNA chains de novo, but are capable of extending the 3' hydroxy terminus of a preexisting 'primer' chain stably annealed to the template strand. The report that partially purified preparations of the dnaG protein, a gene product essential to the replication of E. coli, synthesize RNA primers on phage G4 single-stranded DNA (Bouche, Zechel and Kornberg, 1975) stimulated an investigation into the properties of this enzyme. A thermolabile dnaG protein, prepared from a temperature-sensitive strain of E. coli, was utilized to demonstrate that the ability to prime DNA synthelsis on phage G4 and 0x174 single-stranded DNA resides with the dnaG protein, and that the priming event may be separated from subsequent DNA synthesis. Priming on G4 DNA absolutely requires the E. coli DNA binding protein.
    [Show full text]
  • Coulombic Interactions in Ribonuclease A
    COULOMBIC INTERACTIONS IN RIBONUCLEASE A by Barbra Mindy Templer A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy (Biochemistry) at the UNIVERSITY OF WISCONSIN-MADISON 1998 A dissertation entitled Coulombic Interactions in Ribonuclease A submitted to the Graduate School of the University of Wisconsin-Madison in partial fulfillment of the requirements for the degree of Doctor of Philosophy by Barbra Mindy Templer Date of Final Oral Examination: May 21, 1998 1998 Month & Year Degree to be awarded: December May August * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * • * * * * * * * * * * * * * * * * * * * * * * Approval Signal ures of Dissertation Readers: Signature, Dean of Graduate School l---[-;:-~ ,,(1~ h~~~ ~~~ ~ /tYk ,ec,~~ COULOMBIC INTERACTIONS IN RIBONUCLEASE A Barbra Mindy Templer Under the supervision of Associate Professor Ronald T. Raines At the University of Wisconsin-Madison The interactions between bovine pancreatic ribonuclease (RNase) A and its RNA substrate extend beyond the scissile P-Os' bond. Enzymic subsites interact with the bases and phosphoryl groups of the substrate. Those residues interacting with the phosphoryl groups comprise the PO, PI, and P2 subsites, with the scissile bond residing in the PI subsite. In this Dissertation, the function of the PO and P2 subsites of RNase A is characterized in detail. A new subsite, P( -1), is also described. Lys66 (PO subsite) and Lys7 and Argl0 (P2 subsite) were replaced with alanine residues. Wild-type RNase A and the K66A, K7 AJRlOA, and K7 AJRlONK66A variants were evaluated as catalysts for the cleavage of poly(cytidylic acid) and for their abilities to bind to single-stranded DNA, a substrate analog (Chapter Two). The values of kcal' Km, and kcalKm for poly(C) cleavage were affected by altering the PO and P2 subsites as were the values of Kd for RNase A-d(AUAA.) complex formation.
    [Show full text]
  • Quenching-Resolved Emission Anisotropy Studies with Single and Multitryptophan-Containing Proteins
    QUENCHING-RESOLVED EMISSION ANISOTROPY STUDIES WITH SINGLE AND MULTITRYPTOPHAN-CONTAINING PROTEINS MAURICE EFTINK Department ofChemistry, University ofMississippi, University, Mississippi 38677 ABSTRACT Measurements of the anisotropy ofprotein fluorescence as a function of an added collisional quencher, such as acrylamide, are used to construct Perrin plots. For single tryptophan containing proteins, such plots yield an apparent rotational correlation time for the depolarization process, which, in most cases, is approximately the value expected for Brownian rotation of the entire protein. Apparent limiting fluorescence anisotropy values, which range from 0.20 to 0.32 for the proteins studied, are also obtained from the Perrin plots. The lower values for the limiting anisotropy found for some proteins are interpreted as indicating the existence of relatively rapid, limited (within a cone of angle 0°-30°) motion of the trytophan side chains that is independent of the overall rotation of the protein. Examples of the use of this fluorescence technique to study protein conformational changes are presented, including the monomer - dimer equilibrium of P-lactoglobulin, the monomer - tetramer equilibrium of melittin, the N - F transition of human serum albumin, and the induced change in the conformation of cod parvalbumin caused by the removal of Ca+2. Because multitryptophan-containing proteins have certain tryptophans that are accessible to solute quencher and others that are inaccessible, this method can be used to determine the steady state anisotropy of each class of tryptophan residues. INTRODUCTION varying temperature or the bulk viscosity (see discussion in Studies of the anisotropy of the fluorescence of trypto- Concluding Remarks). phanyl residues in proteins can provide valuable informa- Another steady state approach, used first by Teale and tion concerning the dynamics of these macromolecular Badley (16), involves the measurement of the emission structures.
    [Show full text]
  • The Development of the Prediction of Protein Structure
    6 The Development of the Prediction of Protein Structure Gerald D. Fasman I. Introduction .................................................................... 194 II. Protein Topology. .. 196 III. Techniques of Protein Prediction ................................................... 198 A. Sequence Alignment .......................................................... 199 B. Hydrophobicity .............................................................. 200 C. Minimum Energy Calculations ................................................. 202 IV. Approaches to Protein Conformation ................................................ 203 A. Solvent Accessibility ......................................................... 203 B. Packing of Residues .......................................................... 204 C. Distance Geometry ........................................................... 205 D. Amino Acid Physicochemical Properties ......................................... 205 V. Prediction of the Secondary Structure of Proteins: a Helix, ~ Strands, and ~ Turn .......... 208 A. ~ Turns .................................................................... 209 B. Evaluation of Predictive Methodologies .......................................... 218 C. Other Predictive Algorithms ................................................... 222 D. Chou-Fasman Algorithm ...................................................... 224 E. Class Prediction ............................................................. 233 VI. Prediction of Tertiary Structure ...................................................
    [Show full text]
  • Wo 2008/127291 A2
    (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (43) International Publication Date PCT (10) International Publication Number 23 October 2008 (23.10.2008) WO 2008/127291 A2 (51) International Patent Classification: Jeffrey, J. [US/US]; 106 Glenview Drive, Los Alamos, GOlN 33/53 (2006.01) GOlN 33/68 (2006.01) NM 87544 (US). HARRIS, Michael, N. [US/US]; 295 GOlN 21/76 (2006.01) GOlN 23/223 (2006.01) Kilby Avenue, Los Alamos, NM 87544 (US). BURRELL, Anthony, K. [NZ/US]; 2431 Canyon Glen, Los Alamos, (21) International Application Number: NM 87544 (US). PCT/US2007/021888 (74) Agents: COTTRELL, Bruce, H. et al.; Los Alamos (22) International Filing Date: 10 October 2007 (10.10.2007) National Laboratory, LGTP, MS A187, Los Alamos, NM 87545 (US). (25) Filing Language: English (81) Designated States (unless otherwise indicated, for every (26) Publication Language: English kind of national protection available): AE, AG, AL, AM, AT,AU, AZ, BA, BB, BG, BH, BR, BW, BY,BZ, CA, CH, (30) Priority Data: CN, CO, CR, CU, CZ, DE, DK, DM, DO, DZ, EC, EE, EG, 60/850,594 10 October 2006 (10.10.2006) US ES, FI, GB, GD, GE, GH, GM, GT, HN, HR, HU, ID, IL, IN, IS, JP, KE, KG, KM, KN, KP, KR, KZ, LA, LC, LK, (71) Applicants (for all designated States except US): LOS LR, LS, LT, LU, LY,MA, MD, ME, MG, MK, MN, MW, ALAMOS NATIONAL SECURITY,LLC [US/US]; Los MX, MY, MZ, NA, NG, NI, NO, NZ, OM, PG, PH, PL, Alamos National Laboratory, Lc/ip, Ms A187, Los Alamos, PT, RO, RS, RU, SC, SD, SE, SG, SK, SL, SM, SV, SY, NM 87545 (US).
    [Show full text]
  • United States Patent (19) 11 Patent Number: 6,103,881 Nett Et Al
    USOO6103881A United States Patent (19) 11 Patent Number: 6,103,881 Nett et al. (45) Date of Patent: Aug. 15, 2000 54) GNRH ANALOGS FOR DESTROYING Chang et al., “Artificial Hybrid Protein Containing a Toxin GONADOTROPHS Protein Fragment and a Cell Membrane Receptor-Binding Moiety in a Disulfide Conjugate”, pp. 1515-1522, 1977, J. 75 Inventors: Torrance M. Nett, Ft. Collins; Biol. Chem., vol. 252, Feb. Leonard Michael Glode, Aurora; Chaudhary et al., “Activity of a Recombinant Fusion Protein Marat Karpeisky, Boulder, all of Colo. Between Transforming Growth Factor Type C. and Pseudomonas Toxin'", pp. 4538-4542, 1987, Proc. Natl. 73 Assignee: Colorado State University Research Acad. Sci. USA, Vol. 84, Jul. Foundation, Ft. Collins, Colo. Colombatti et al., “Cloned Fragment of Diphtheria Toxin Linked to T Cell-specific Antibody Identifies Regions of B 21 Appl. No.: 09/015,729 Chain Active in Cell Entry”, pp. 3030-3035, 1986 J. Biol. 22 Filed: Apr. 7, 1998 Chem., vol. 261, Mar. Greenfield et al., “Mutations in Diphtheria Toxin Separate Related U.S. Application Data Binding from Entry and Amplify Immunotoxin Selectivity”, pp. 536-539, 1987, Science, vol. 238, Oct. 63 Continuation-in-part of application No. 07/314,653, Feb. 23, Meyers et al., “Specific Chemical Cleavage of Diphtheria 1989, abandoned, and a division of application No. 08/481, Toxin With Hydroxylamine”, pp. 17122–17127, 1988, J. 128, Jun. 7, 1995, Pat. No. 5,786,457. Biol. Chem., vol. 263, No. 32, Nov. 51 Int. Cl." ............................. A61K38/00; CO7K 1/00; Murphy et al., “Genetic Construction, Expression, and C07K5/00; CO7K 7/00 Melanoma-Selective Cytotoxicity of a Diphtheria Toxin 52 U.S.
    [Show full text]
  • Polyamines Mitigate Antibiotic Inhibition of A.Actinomycetemcomitans Growth
    Polyamines Mitigate Antibiotic Inhibition of A.actinomycetemcomitans Growth THESIS Presented in Partial Fulfillment of the Requirements for the Degree Master of Science in the Graduate School of The Ohio State University By Allan Wattimena Graduate Program in Dentistry The Ohio State University 2017 Master's Examination Committee: Dr John Walters, Advisor Dr Purnima Kumar Dr Sara Palmer Dr Shareef Dabdoub Copyright by Allan Wattimena 2017 Abstract Polyamines are ubiquitous polycationic molecules that are present in all prokaryotic and eukaryotic cells. They are the breakdown products of amino acids and are important modulators of cell growth, stress and cell proliferation. Polyamines are present in higher concentrations in the periodontal pocket and may affect antibiotic resistance of bacterial biofilms. The effect of polyamines was investigated with amoxicillin (AMX), azithromycin (AZM) and doxycycline (DOX) on the growth of Aggregatibacter actinomycetemcomitans (A.a.) Y4 strain. Bacteria were grown in brain heart infusion broth under the following conditions: 1) A.a. only, 2) A.a. + antibiotic, 3) A.a. + antibiotic + polyamine mix (1.4mM putrescine, 0.4mM spermidine, 0.4mM spermine). Growth curve analysis, MIC determination and metatranscriptomic analysis were carried out. The presence of exogenous polyamines produced a small, but significant increase in growth of A.a. Polyamines mitigated the inhibitory effect of AMX, AZM and DOX on A.a. growth. Metatranscriptomic analysis revealed differing transcriptomic profiles when comparing AMX and AZM in the presence of polyamines. Polyamines produced a transient mitigation of AMX inhibition, but did not have a significant effect on gene transcription. Many gene transcription changes were seen when polyamines were in the presence of AZM.
    [Show full text]
  • Introduction the Terms in the Subject Index for Volume 81, January
    Introduction The terms in the Subject Index for Volume 81, January-December 1984, of the PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES USA (Biological Sci- ences) were chosen from titles, key terms, and abstracts of articles. The index terms are alphabetized by computer; numbers, conformational prefixes, and hy- phenated Greek letters are disregarded in alphabetization. 8041 8042 Biological Sciences Subject Index Proc. Natl. Acad. Sci. USA 81 (1984) A* protein Different channel properties of Torpedo acetylcholine receptor monomers Cleavage of single-stranded DNA by the <X174 A* protein: The A*- and dimers reconstituted in planar membranes, 6222 single-stranded DNA covalent linkage, 4285 Phosphorylation of the nicotinic acetylcholine receptor by an endogenous Abelson murine leukemia virus tyrosine-specific protein kinase, 6968 Transformation-associated proteins in murine B-cell lymphomas that are Purification of the muscarinic acetylcholine receptor from porcine atria distinct from Abelson virus gene products, 4434 (Correction), 7258 Abortive infection Acetylcholine receptor-specific suppressive T-cell factor from a Virus-plasmid interactions: Mutants of bacteriophage T3 that abortively retrovirally transformed T-cell line, 7569 infect plasmid F-containing (F+) strains of Escherichia coli, 1465 Isolation and characterization of a cDNA clone for the complete protein Cytomegalovirus infects human lymphocytes and monocytes: Virus coding region of the 8 subunit of the mouse acetylcholine expression is restricted to immediate-early gene products,
    [Show full text]