DOCSLIB.ORG

(Adenine to Cytosine) Resulting in a Failure to Cleave Amino-Terminal Methionine (Globin/Initiation Codon/Methionine Cleavage/Protein Processing) JOSEF T

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
(Adenine to Cytosine) Resulting in a Failure to Cleave Amino-Terminal Methionine (Globin/Initiation Codon/Methionine Cleavage/Protein Processing) JOSEF T Proc. Natl. Acad. Sci. USA Vol. 83, pp. 24-27, January 1986 Biochemistry Hemoglobin Long Island is caused by a single mutation (adenine to cytosine) resulting in a failure to cleave amino-terminal methionine (globin/initiation codon/methionine cleavage/protein processing) JOSEF T. PRCHAL*t, DANIEL P. CASHMANt, AND Y. W. KANt tHoward Hughes Institute and Department of Medicine, University of California, San Francisco, CA 94143; and *Hemoglobin and Red Cell Laboratory and Division of Hematology, University of Alabama at Birmingham, Birmingham, AL 35294 Communicated by Helen M. Ranney, August 26, 1985 ABSTRACT Hemoglobin Long Island has two separate trout cells (11). It is removed from the nascent polypeptide amino acid abnormalities of /3-globin structure: an extension of chain by posttranslational modification in most nonsecreted the NH2 terminus by a methionine residue and a histidine-to- prokaryotic and eukaryotic proteins (12). Exceptions to this proline substitution at the normal second position. The NH2- rule are found in various prokaryotic and eukaryotic proteins terminal methionine residue, the translation product of an (13) and, also perhaps relevant to this report, in some AUG initiation codon, is present only transiently in nascent ruminant p-globins such as bovine f3- and A-related y-globin proteins. Because of the general biological implications of this (14, 15) as well as in other ruminants-i.e., sheep and goat abnormality, we investigated the nature of the genetic defect of (16). this mutant. We determined the sequence of the relevant portion of the f8-globin mRNA by means of dideoxynucleotide MATERIALS AND METHODS chain termination ofthe complementary DNA (cDNA) in which an oligonucleotide complementary to codons 10-17 was used as Subjects and Reticulocyte RNA Preparation. The mutant a primer for reverse transcriptase. A histidine-to-proline hemoglobin was discovered because ofits interference with the substitution was confirmed in the mutant mRNA by identifying detection of glycosylated hemoglobin in the propositus who is an adenine-to-cytosine transversion in the second codon. How- a diabetic, and the same mutant was also subsequently found in ever, we were unable to find any other abnormality at either the his nondiabetic mother. No apparent deleterious effect was AUG initiation codon or in the 56 bases upstream from the associated with the Hb Long Island in either the propositus or adenine-to-cytosine transversion (encompassing most of the 5' his mother. Heparinized blood from the propositus and his untranslated region of the mutant (3-globin mRNA). Thus, it mother (both heterozygous for Hb Long Island) and from a appears that this single lesion probably interferes with the control subject undergoing an exchange transfusion for pro- poorly understood methionine-cleaving mechanism that mod- longed sickle-cell crisis served as the source of globin mRNA. ulates most of prokaryotic and eukaryotic proteins. Erythrocytes were selectively lysed by exposure to 0.1444 M NH4Cl/0.001 M NH4HCO3 solution, and crude RNA was We have recently described a hemoglobin mutant, hemoglo- precipitated by acid titration. RNA was further purified by bin Long Island (1), that is characterized by two separate phenol/chloroform, 50:50 (vol/vol) extraction and ethanol pre- abnormalities in the primary amino acid structure of the cipitation (17). Neither ofthe two subjects with Hb Long Island P3-globin chain: (i) an extension of the NH2 terminus by a had evidence of shortened erythrocyte survival; thus, only 30 methionine residue and (ii) the substitution of a proline for a tkg (mother) and 34 tkg (propositus) of total reticulocyte RNA histidine at the second amino acid position. A similar mutant were obtained from each 100 ml peripheral blood. has also been described in France as hemoglobin Marseille Oligonucleotide Primer Synthesis. The deoxyoligonucleo- (2). Although more than 400 hemoglobin mutants have been tide primer was 24 nucleotides long and was complementary described (3), this P-globin is ofinterest for two reasons: first, to P-globin codons 10-17. It was constructed on a DNA the extension of the NH2 terminus and, second, of more synthesizer (Applied Biosystems, model 380A, Foster City, general biological significance, the fact that it is a mutant CA) utilizing the phosporamidite (18, 19) method with con- nonsecreted vertebrate protein in which the NH2-terminal trolled-pore-glass solid-phase support and was further puri- methionine, coded by the AUG initiation codon, is preserved fied on an HPLC (IBM, model LC 9533, San Jose, CA) in the protein structure. equipped with a C4 reverse-phase column. Its composition To establish the molecular lesion responsible for this was as follows: dCACCTTGCCCCACAGGGCAGTAAC (5' mutation, we have directly sequenced mRNA from the to 3' orientation). normal and mutant P-globin mRNA of two subjects cDNA Synthesis. The following conditions were established heterozygous for Hb Long Island. The knowledge of the to be optimal for full-length transcription of p-globin cDNA. exact nature of the mutation associated with Hb Long Island 360 pmol of oligonucleotide was mixed with 5 ,ug of total is of interest since the protein abnormality could be caused reticulocyte RNA, heated for 3 min at 98°C, rapidly cooled, by: (i) a general abnormality ofthe poorly defined methionine suspended in annealing buffer [0.4 M NaCl, Pipes (40 mM, cleavage pathway, (ii) two separate mutations, or (iii) a single pH 6.4), 80% (vol/vol) formamide] for 3 hr at 300C, and RNA mutation that interferes with the cleavage of the initial NH2 was ethanol precipitated. Each reaction mixture consisted of terminus. This NH2-terminal methionine has been found to RNA (0.2 ,ug in 2-,u aliquots) suspended in a solution of be transiently present in nascent bacterial polypeptides (4), RNasin (0.2 unit/ml; Promega Biotec, Madison, WI), acti- human tumor cells in ascites fluid (5), globins (6-10), and nomycin D, and reverse transcriptase buffer (see below). This RNA solution was incubated in a total volume of 5 ,ul containing 800 units of avian myeloblastosis virus reverse The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact. tTo whom reprint requests should be addressed. 24 Biochemistry: Prchal et al. Proc. Natl. Acad. Sci. USA 83 (1986) 25 transcriptase per ml, Tris HCl (100 mM, pH 8.3), 10 mM propositus heterozygous for Hb Long Island, one with CAC MgCl2, 50 mM KCl, 10 mM dithiothreitol, actinomycin D (40 coding for histidine in the second B-globin codon position and jug/ml), and various amounts of deoxynucleotides with the other with a CCC codon corresponding to proline. Also in approximately 1% of the dCTP labeled with 32p. this gel the adenine-to-thymine substitution at codon 6 resulting ,t-Globin mRNA Sequencing. Nucleotide sequences of the in a glutamine-to-valine substitution (sickle-cell mutation) is desired 13-globin anticodon region of interest in the mutant seen clearly in the control reticulocyte RNA obtained from a and control sickle-cell samples were determined by the patient with sickle-cell disease (Fig. 1). No other abnormality dideoxy method (20). The following concentrations were was seen in this analysis. Results ofthe 12% and 8% acrylamide found to be optimal for the sequencing reactions: gels confirmed the adenine-to-cytosine transversion. Separate deoxynucleotides (dCTP, 1.1 ,uM; dATP, dGTP, and dTTP, sequencing of RNA from the propositus' mother revealed the 26 AuM) and dideoxynucleotides (ddCTP, 0.4 ,uM; ddATP, 40 identical abnormality to that of the propositus. ,M; ddGTP, 8 ,uM; ddTTP, 20 ,uM). After 60 min at 37°C, each reaction was stopped by adding 4 ,ul of a solution containing 0.03% xylene cyanol FF, 0.03% bromphenol blue, DISCUSSION EDTA (10 mM, pH 8.3), 90%o (vol/vol) formamide and heated To our knowledge this is the first described naturally occur- at 65°C for 15 min. These mixtures were then rapidly cooled ring protein mutant characterized by NH2-terminal methio- and applied in 3-,ul aliquots to acrylamide gels containing 7 M nine extension. The analysis of the mRNA sequence of the urea. To increase the resolution of the various mRNA /3-globin mutant, Hb Long Island, confirmed the histidine- regions, 8%, 12%, and 20% acrylamide gels were used and to-proline substitution (1) resulting from an adenine-to- analyzed by autoradiography. cytosine transversion. The RNA sequence immediately ad- jacent to the AUG initiation codon was normal, as was the RESULTS more upstream region of the untranslated portion of 83-globin mRNA (21). Although the mutant protein consisted of two An essentially full-length cDNA transcript (better than 95% separate amino acid abnormalities (1), no other nucleic acid complete) was obtained at concentrations of deoxynucleotides mutation was found. greater than 50 ,uM. This transcript of the translated and 5' We have determined the nature of the mutation leading to untranslated regions of 8-globin mRNA consisted of 107 bases two separate amino acid abnormalities in the P-globin mutant (data not shown). Even when concentrations of deoxynucleo- of Hb Long Island by direct, rapid sequencing ofthe mRNA. tides optimal for sequencing were used, most of the in vitro This method bypasses the customary, time-consuming DNA synthesized cDNA was completely transcribed. Sequence anal- cloning and sequencing methods. RNA sequencing of homo- ysis of the reticulocyte mRNA derived from the carriers of Hb geneous viral RNA by the dideoxynucleotide termination Long Island showed that both the CAC and CCC sequences method has been accomplished (23). Eukaryotic mRNA were present at codon number two (Fig. 1). This confirmed the [poly(A)-rich RNA] has also been sequenced by this method histidine-to-proline substitution (1) in this position (Fig.
Load more

Recommended publications
  • Effects of Single Amino Acid Deficiency on Mrna Translation Are Markedly
    www.nature.com/scientificreports OPEN Efects of single amino acid defciency on mRNA translation are markedly diferent for methionine Received: 12 December 2016 Accepted: 4 May 2018 versus leucine Published: xx xx xxxx Kevin M. Mazor, Leiming Dong, Yuanhui Mao, Robert V. Swanda, Shu-Bing Qian & Martha H. Stipanuk Although amino acids are known regulators of translation, the unique contributions of specifc amino acids are not well understood. We compared efects of culturing HEK293T cells in medium lacking either leucine, methionine, histidine, or arginine on eIF2 and 4EBP1 phosphorylation and measures of mRNA translation. Methionine starvation caused the most drastic decrease in translation as assessed by polysome formation, ribosome profling, and a measure of protein synthesis (puromycin-labeled polypeptides) but had no signifcant efect on eIF2 phosphorylation, 4EBP1 hyperphosphorylation or 4EBP1 binding to eIF4E. Leucine starvation suppressed polysome formation and was the only tested condition that caused a signifcant decrease in 4EBP1 phosphorylation or increase in 4EBP1 binding to eIF4E, but efects of leucine starvation were not replicated by overexpressing nonphosphorylatable 4EBP1. This suggests the binding of 4EBP1 to eIF4E may not by itself explain the suppression of mRNA translation under conditions of leucine starvation. Ribosome profling suggested that leucine deprivation may primarily inhibit ribosome loading, whereas methionine deprivation may primarily impair start site recognition. These data underscore our lack of a full
    [Show full text]
  • Enzymatic Assay of L-METHIONINE GAMMA-LYASE (EC 4.4.1.11)
    Enzymatic Assay of L-METHIONINE GAMMA-LYASE (EC 4.4.1.11) PRINCIPLE: L-Methionine Gamma-Lyase L-Methionine > Methanethiol + 2-Ketobutyrate + NH3 2-Ketobutyrate + MBTH > Azine Derivative Abbreviation used: MBTH = 3-Methyl-2-Benzothiazolinone CONDITIONS: T = 37°C, pH = 8.0, A320nm, Light path = 1 cm METHOD: Stopped Spectrophotometric Rate Determination REAGENTS: A. 100 mM Potassium Phosphate Buffer with 25 mM L-Methionine and 0.01 mM Pyridoxal 5-Phosphate, pH 8.0 at 37°C1 (Prepare 100 ml in deionized water using Potassium Phosphate, Monobasic, Anhydrous, Sigma Prod. No. P-5379, L-Methionine, Sigma Prod. No. M-9625, and Pyridoxal 5-Phosphate, Sigma Prod. No. P-9255. Adjust to pH 8.0 at 37°C with 5 M KOH.) B. 50% (w/v) Trichloroacetic Acid Solution (TCA) (Prepare 5 ml in deionized water using Trichloroacetic Acid, 6.1 N Solution, approximately 100% (w/v), Sigma Stock No. 490-10.) C. 1 M Sodium Acetate Buffer, pH 5.0 at 37°C (NaOAC) (Prepare 100 ml in deionized water using Sodium Acetate, Trihydrate, Sigma Prod. No. S-8625. Adjust to pH 5.0 at 37°C with 5 M HCl.) D. 0.1% (w/v) 3-Methyl-2-Benzothiazolinone Hydrazone (MBTH) (Prepare 10 ml in deionized water using 3-Methyl-2-Benzothiazolinone Hydrazone, Hydrochloride Hydrate, Sigma Prod. No. M-8006.) SSMETH01 Page 1 of 4 07/29/98 Enzymatic Assay of L-METHIONINE GAMMA-LYASE (EC 4.4.1.11) REAGENTS: E. 100 mM Potassium Phosphate Buffer with 1 mM Ethylenediaminetetraacetic Acid, 0.01% (v/v) 2-Mercaptoethanol and 0.02 mM Pyridoxal 5-Phosphate, pH 7.2 at 37°C (Enzyme Diluent)1 (Prepare 10 ml in deionized water using Potassium Phosphate, Monobasic, Anhydrous, Sigma Prod.
    [Show full text]
  • Relative Reaction Rates of the Amino Acids Cysteine, Methionine, and Histidine with Analogs of the Anti-Cancer Drug Cisplatin Cynthia A
    Western Kentucky University TopSCHOLAR® Honors College Capstone Experience/Thesis Honors College at WKU Projects 5-11-2015 Relative Reaction Rates of the Amino Acids Cysteine, Methionine, and Histidine with Analogs of the Anti-Cancer Drug Cisplatin Cynthia A. Tope Western Kentucky University, [email protected] Follow this and additional works at: http://digitalcommons.wku.edu/stu_hon_theses Part of the Medicinal-Pharmaceutical Chemistry Commons Recommended Citation Tope, Cynthia A., "Relative Reaction Rates of the Amino Acids Cysteine, Methionine, and Histidine with Analogs of the Anti-Cancer Drug Cisplatin" (2015). Honors College Capstone Experience/Thesis Projects. Paper 571. http://digitalcommons.wku.edu/stu_hon_theses/571 This Thesis is brought to you for free and open access by TopSCHOLAR®. It has been accepted for inclusion in Honors College Capstone Experience/ Thesis Projects by an authorized administrator of TopSCHOLAR®. For more information, please contact [email protected]. RELATIVE REACTION RATES OF THE AMINO ACIDS CYSTEINE, METHIONINE, AND HISTIDINE WITH ANALOGS OF THE ANTI-CANCER DRUG CISPLATIN A Capstone Experience/Thesis Project Presented in Partial Fulfillment of the Requirements for the Degree Bachelor of Science with Honors College Graduate Distinction at Western Kentucky University By: Cynthia A. Tope ***** Western Kentucky University 2015 CE/T Committee: Approved by: Professor Kevin Williams, Advisor _________________________ Professor Darwin Dahl Advisor Professor Lee Ann Smith Department of Chemistry Copyright: Cynthia A. Tope 2015 ABSTRACT We are studying the reaction of analogs of the anticancer drug cisplatin with amino acids that differ in size and shape. The reaction of cisplatin with proteins likely precedes reaction with DNA in the body, forming a variety of products that may be toxic to the human body.
    [Show full text]
  • Lyxumia Subcutaneous Injection 300 Μg [Non-Proprietary Name] Lixisenatide (JAN*) [Name of Applicant] Sanofi K.K
    Report on the Deliberation Results May 31, 2013 Evaluation and Licensing Division, Pharmaceutical and Food Safety Bureau Ministry of Health, Labour and Welfare [Brand name] Lyxumia Subcutaneous Injection 300 μg [Non-proprietary name] Lixisenatide (JAN*) [Name of applicant] Sanofi K.K. [Date of application] June 11, 2012 [Results of deliberation] In the meeting held on May 24, 2013, the First Committee on New Drugs concluded that the product may be approved and that this result should be reported to the Pharmaceutical Affairs Department of the Pharmaceutical Affairs and Food Sanitation Council. The re-examination period for the product is 8 years, and the drug substance and the drug product are both classified as powerful drugs and the product is not classified as a biological product or a specified biological product. The proposed Japanese brand name should be changed for ensuring medical safety. *Japanese Accepted Name (modified INN) This English version of the Japanese review report is intended to be a reference material to provide convenience for users. In the event of inconsistency between the Japanese original and this English translation, the former shall prevail. The PMDA will not be responsible for any consequence resulting from the use of this English version. Review Report May 7, 2013 Pharmaceuticals and Medical Devices Agency The results of a regulatory review conducted by the Pharmaceuticals and Medical Devices Agency on the following pharmaceutical product submitted for registration are as follows. [Brand name] Lyxumia Subcutaneous
    [Show full text]
  • BASIC GENETICS for the CLINICAL NEUROLOGIST M R Placzek, T T Warner *Ii5
    J Neurol Neurosurg Psychiatry: first published as 10.1136/jnnp.73.suppl_2.ii5 on 1 December 2002. Downloaded from BASIC GENETICS FOR THE CLINICAL NEUROLOGIST M R Placzek, T T Warner *ii5 J Neurol Neurosurg Psychiatry 2002;73(Suppl II):ii5–ii11 o the casual observer, the clinical neurologist and molecular geneticist would appear very dif- ferent species. On closer inspection, however, they actually have a number of similarities: they Tboth use a rather impenetrable language littered with abbreviations, publish profusely with- out seeming to alter the course of clinical medicine, and make up a small clique regarded as rather esoteric by their peers. In reality, they are both relatively simple creatures who rely on basic sets of rules to work in their specialities. Indeed they have had a productive symbiotic relationship in recent years and the application of molecular biological techniques to clinical neuroscience has had a profound impact on the understanding of the pathophysiology of many neurological diseases. One reason for this is that around one third of recognisable mendelian disease traits demonstrate phenotypic expression in the nervous system. The purpose of this article is to demystify the basic rules of molecular biology, and allow the clinical neurologist to gain a better understanding of the techniques which have led to the isola- tion (cloning) of neurological disease genes and the potential uses of this knowledge. c NUCLEIC ACIDS Deoxyribonucleic acid (DNA) is the macromolecule that stores the blueprint for all the proteins of the body. It is responsible for development and physical appearance, and controls every biological copyright.
    [Show full text]
  • SATMF Suppresses the Premature Senescence Phenotype of the ATM Loss-Of-Function Mutant and Improves Its Fertility in Arabidopsis
    Supplementary Materials SATMF Suppresses the Premature Senescence Phenotype of the ATM Loss-of-Function Mutant and Improves Its Fertility in Arabidopsis Yi Zhang, Hou-Ling Wang, Yuhan Gao, Hongwei Guo and Zhonghai Li Figure S1. Histological GUS staining analysis of gene expression of ATM at different developmental stages. (A) Rosette leaves of 8-day-old seedling of pATM-GUS/Col-0. (B) Rosette leaves of 16-day-old plant of pATM-GUS/Col-0. (C) Rosette leaves of 48-day-old plant of pATM-GUS/Col-0. Int. J. Mol. Sci. 2020, 21, x; doi: FOR PEER REVIEW www.mdpi.com/journal/ijms Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 2 of 5 Figure S2. Screen for suppressor of atm mutant in fertility (satmf) by EMS. (A) Experimental design for screening the expected phenotypes of stamf mutants. (B) EMS mutagenesis of atm-2 seeds by using different experimental conditions. (C) Seeds of atm-2 mutant is hypersensitive to EMS in comparison to Col-0. Int. J. Mol. Sci. 2020, 21, x FOR PEER REVIEW 3 of 5 Figure S3. Molecular identification of satmf mutants. (A) Fertility of plants of Col-0, atm-2, satmf1, satmf2 and satmf3. (B) Genotyping analysis of atm-2, satmf1, satmf2 and satmf3 plants. (C) Confirmation of the backgrounds of satmf1, satmf2 and satmf3 plants by sequencing analysis of PCR products in (B). The “G” PCR reaction tests for the ability to amplify a genome region that will be present in wild type and heterozygous lines, but will not amplify in homozygous lines.
    [Show full text]
  • The Un-Design and Design of Insulin: Structural Evolution
    THE UN-DESIGN AND DESIGN OF INSULIN: STRUCTURAL EVOLUTION WITH APPLICATION TO THERAPEUTIC DESIGN By NISCHAY K. REGE Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy Department of Biochemistry Dissertation Advisor: Dr. Michael A. Weiss CASE WESTERN RESERVE UNIVERSITY August, 2018 CASE WESTERN RESERVE UNIVERSITY SCHOOL OF GRADUATE STUDIES We hereby approve the thesis/dissertation of Nischay K. Rege candidate for the degree of Doctor of Philosophy*. Committee Chair Paul Carey Committee Member Michael Weiss Committee Member Faramarz Ismail-Beigi Committee Member George Dubyak Date of Defense: June 26th, 2018 *We also certify that written approval has been obtained for any proprietary material contained therein. Dedication This thesis is dedicated to my mother, Dipti, whose constant love and faith have never failed, to my father, Kiran, who taught me of the virtue of curiosity, and to my wife, Shipra, whose kindness and companionship have given me enough strength for eight lifetimes. i Table of Contents Dedication ..................................................................................................................................... i Table of Contents ......................................................................................................................... ii List of Tables ............................................................................................................................... v List of Figures ...........................................................................................................................
    [Show full text]
  • Genetic Studies of Leptin Concentrations Implicate Leptin in the Regulation of Early
    Page 1 of 93 Diabetes Genetic studies of leptin concentrations implicate leptin in the regulation of early adiposity Hanieh Yaghootkar1,2,3*&, Yiying Zhang4&, Cassandra N Spracklen5, Tugce Karaderi6,7,8,9, Lam Opal Huang10, Jonathan Bradfield11,12, Claudia Schurmann13, Rebecca S Fine14,15,16, Michael H Preuss13, Zoltan Kutalik1,17,18, Laura BL Wittemans6,19, Yingchang Lu13,20, Sophia Metz10, Sara M Willems19, Ruifang Li-Gao21, Niels Grarup10, Shuai Wang22, Sophie Molnos23,24, América A Sandoval-Zárate25, Mike A Nalls26,27, Leslie A Lange28, Jeffrey Haesser29, Xiuqing Guo30, Leo-Pekka Lyytikäinen31,32, Mary F Feitosa33, Colleen M Sitlani34, Cristina Venturini35, Anubha Mahajan6,36, Tim Kacprowski37,38, Carol A Wang22, Daniel I Chasman39,40, Najaf Amin41, Linda Broer42, Neil Robertson6,36, Kristin L Young43, Matthew Allison44, Paul L Auer45, Matthias Blüher46, Judith B Borja47,48, Jette Bork-Jensen10, Germán D Carrasquilla10, Paraskevi Christofidou35, Ayse Demirkan41, Claudia A Doege49, Melissa E Garcia50, Mariaelisa Graff43,51, Kaiying Guo4, Hakon Hakonarson11,52, Jaeyoung Hong22, Yii-Der Ida Chen30, Rebecca Jackson53, Hermina Jakupović10, Pekka Jousilahti54, Anne E Justice55, Mika Kähönen56,57, Jorge R Kizer58,59, Jennifer Kriebel23,24, Charles A LeDuc4, Jin Li60, Lars Lind61, Jian’an Luan19, David Mackey62, Massimo Mangino35,63, Satu Männistö54, Jayne F Martin Carli4, Carolina Medina-Gomez41,42, Dennis O Mook-Kanamori21,64, Andrew P Morris65,6,66, Renée de Mutsert21, Matthias Nauck67,38, Ivana Nedeljkovic41, Craig E Pennell22, Arund D Pradhan39,40, Bruce M Psaty68,69, Olli T Raitakari70,71,72, Robert A Scott19, Tea Skaaby73, Konstantin Strauch74,75, Kent D Taylor30, Alexander Teumer76,38, Andre G Uitterlinden41,42, Ying Wu5, Jie Yao30, Mark Walker77, Kari E North43, Peter Kovacs46, M.
    [Show full text]
  • L-Methionine
    October 24, 2011 Lisa Brines, Ph.D. National List Manager USDA/AMS/NOP, Standards Division 1400 Independence Ave. SW Room 2646-So., Ag Stop 0268 Washington, DC 20250-0268 RE: Petition for inclusion of L-Methionine on the National List at §205.605(b) as a synthetic non-agricultural substance allowed in or on processed infant formula products labeled as “organic” or “made with organic (specified ingredients)” with the annotation “for use only in infant formula based on isolated soy protein.” Dear Dr. Brines, The International Formula Council (IFC) is an association of manufacturers and marketers of formulated nutrition products (e.g., infant formulas and adult nutritionals) whose members are based predominantly in North America. IFC members support the American Academy of Pediatrics’ (AAP) position that breastfeeding is the preferred method of feeding infants. We also agree with the AAP that, for infants who do not receive breast milk, iron-fortified infant formula is the only safe and recommended alternative, IFC members are committed to providing infant formulas of the highest quality for those mothers who cannot or choose not to breastfeed, discontinue breastfeeding prior to one year or choose to supplement. This petition seeks to add L-Methionine to the National List to permit its addition as a nonagricultural ingredient in infant formula based on isolated soy protein. L-Methionine is an essential amino acid for humans of all ages. Amino acids are the building blocks of protein. An essential amino acid is one that must be provided in the foods in our diet since our bodies do not have the capability of producing enough of it for normal metabolism and growth.
    [Show full text]
  • Amino Acid Transport Pathways in the Small Intestine of the Neonatal Rat
    Pediat. Res. 6: 713-719 (1972) Amino acid neonate intestine transport, amino acid Amino Acid Transport Pathways in the Small Intestine of the Neonatal Rat J. F. FITZGERALD1431, S. REISER, AND P. A. CHRISTIANSEN Departments of Pediatrics, Medicine, and Biochemistry, and Gastrointestinal Research Laboratory, Indiana University School of Medicine and Veterans Administration Hospital, Indianapolis, Indiana, USA Extract The activity of amino acid transport pathways in the small intestine of the 2-day-old rat was investigated. Transport was determined by measuring the uptake of 1 mM con- centrations of various amino acids by intestinal segments after a 5- or 10-min incuba- tion and it was expressed as intracellular accumulation. The neutral amino acid transport pathway was well developed with intracellular accumulation values for leucine, isoleucine, valine, methionine, tryptophan, phenyl- alanine, tyrosine, and alanine ranging from 3.9-5.6 mM/5 min. The intracellular accumulation of the hydroxy-containing neutral amino acids threonine (essential) and serine (nonessential) were 2.7 mM/5 min, a value significantly lower than those of the other neutral amino acids. The accumulation of histidine was also well below the level for the other neutral amino acids (1.9 mM/5 min). The basic amino acid transport pathway was also operational with accumulation values for lysine, arginine and ornithine ranging from 1.7-2.0 mM/5 min. Accumulation of the essential amino acid lysine was not statistically different from that of nonessential ornithine. Ac- cumulation of aspartic and glutamic acid was only 0.24-0.28 mM/5 min indicating a very low activity of the acidic amino acid transport pathway.
    [Show full text]
  • OBSERVATIONS Sured, As Indicated in Fig
    LETTERS blood glucose concentrations were mea- lispro at 40 Ϯ 3 min and aspart at 49 Ϯ 3 OBSERVATIONS sured, as indicated in Fig. 1. If blood glu- min after injection, respectively (P ϭ cose was 3.5 mmol/l or lower, 20 ml 0.01). The maximum insulin concentra- glucose 30% was injected. One patient tion was 316 Ϯ 31 pmol/l on insulin lis- Direct Comparison of was excluded from the analysis because, pro and 295 Ϯ 27 pmol/l on insulin by mistake, he took a large extra dose of aspart (NS) (Fig. 1). The increase from 0 Insulin Lispro and insulin the night before the study. to 15 min after injection was 109 Ϯ 17 Aspart Shows Small Free insulin was measured after poly- pmol/l after injection of insulin lispro and Differences in ethylene glycol precipitation by Mercodia 53 Ϯ 11 pmol/l after injection of insulin Iso-Insulin (ELISA; Mercodia AB, Uppsala, Plasma Insulin aspart (P ϭ 0.02). Fasting insulin lispro Sweden), a two-site enzyme immunoassay levels reached 50% of peak concentration Profiles After containing two monoclonal antibodies at 20 Ϯ 1 min and aspart at 30 Ϯ 3 min against insulin. Identical results were ob- (P ϭ 0.02) (Fig. 2). The decrease of free Subcutaneous tained when equimolar concentrations of Injection in Type 1 insulin concentration from peak concen- human insulin, insulin lispro, and insulin tration to 50% of the maximum concen- aspart were tested, indicating 100% Diabetes tration was found at 113 Ϯ 10 min during cross-reactivity between lispro, aspart, insulin lispro and 154 Ϯ 14 min during and human insulin in this assay.
    [Show full text]
  • Molecular and Supramolecular Structure of the Mitochondrial Oxidative Phosphorylation System: Implications for Pathology
    life Review Molecular and Supramolecular Structure of the Mitochondrial Oxidative Phosphorylation System: Implications for Pathology Salvatore Nesci 1,* , Fabiana Trombetti 1, Alessandra Pagliarani 1,*, Vittoria Ventrella 1, Cristina Algieri 1 , Gaia Tioli 2 and Giorgio Lenaz 2,* 1 Department of Veterinary Medical Sciences, Alma Mater Studiorum University of Bologna, 40064 Ozzano Emilia, Italy; [email protected] (F.T.); [email protected] (V.V.); [email protected] (C.A.) 2 Department of Biomedical and Neuromotor Sciences, Alma Mater Studiorum University of Bologna, 40138 Bologna, Italy; [email protected] * Correspondence: [email protected] (S.N.); [email protected] (A.P.); [email protected] (G.L.) Abstract: Under aerobic conditions, mitochondrial oxidative phosphorylation (OXPHOS) converts the energy released by nutrient oxidation into ATP, the currency of living organisms. The whole biochemical machinery is hosted by the inner mitochondrial membrane (mtIM) where the protonmo- tive force built by respiratory complexes, dynamically assembled as super-complexes, allows the F1FO-ATP synthase to make ATP from ADP + Pi. Recently mitochondria emerged not only as cell powerhouses, but also as signaling hubs by way of reactive oxygen species (ROS) production. How- ever, when ROS removal systems and/or OXPHOS constituents are defective, the physiological ROS generation can cause ROS imbalance and oxidative stress, which in turn damages cell components. Citation: Nesci, S.; Trombetti, F.; Moreover, the morphology of mitochondria rules cell fate and the formation of the mitochondrial Pagliarani, A.; Ventrella, V.; Algieri, permeability transition pore in the mtIM, which, most likely with the F F -ATP synthase contribu- C.; Tioli, G.; Lenaz, G.
    [Show full text]