DOCSLIB.ORG

Serves As High-Affinity Substrate for Tyrosylprotein Sulfotransferase

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Serves As High-Affinity Substrate for Tyrosylprotein Sulfotransferase Proc. Nati. Acad. Sci. USA Vol. 82, pp. 6143-6147, September 1985 Cell Biology (Glu62,Ala30,Tyr8). serves as high-affinity substrate for tyrosylprotein sulfotransferase: A Golgi enzyme (tyrosine sulfation/protein sorting/chromaffin granules/tyrosine phosphorylation/tubulin) RAYMOND W. H. LEE AND WIELAND B. HUTTNER* Department of Neurochemistry, Max-Planck-Institute for Psychiatry, 8033 Martinsried, Federal Republic of Germany Communicated by Fritz Lipmann, May 10, 1985 ABSTRACT Tyrosylprotein sulfotransferase, the enzyme as substrate are as follows: (i) the only sulfatable hydroxyl catalyzing the sulfation of proteins on tyrosine residues, was group in Glu,Ala,Tyr is on tyrosine, in contrast to physio- characterized by using the acidic polymer containing tyrosine logical protein substrates, which may also contain sulfatable (Glu62,Ala3',Tyr8). (referred to as Glu,Ala,Tyr) as exogenous carbohydrate, serine, and threonine residues; (ii) "protein" substrate. After subcellular fractionation ofa bovine Glu,Ala,Tyr is available in completely unsulfated form in adrenal medulla homogenate, tyrosylprotein sulfotransferase sufficient quantities for enzymological studies, whereas activity was found to be highest in fractions enriched in Golgi physiologically sulfated proteins purified from bi6logical membrane vesicles. Tyrosylprotein sulfotransferase required sources need to be desulfated before being used as sub- the presence of a nonionic detergent for sulfation of exogenous strates; (iii) Glu,Ala,Tyr is likely to contain tyrosine residues Glu,Ala,Tyr, indicating an orientation of the catalytic site of in the vicinity ofacidic amino acid residues. A comparison of the enzyme toward the Golgi lumen. Tyrosylprotein the amino acid sequences surrounding the sulfated tyrosine sulfotransferase was solubilized by Triton X-100, suggesting residues in gastrin (10), cholecystokinin (11), fibrinopeptide that the enzyme was tightly associated with the Golgi mem- B (12), and hirudin (13) shows that these sequences always brane, possibly as an integral membrane protein. The apparent contain acidic amino acids. Using Glu,Ala,Tyr as substrate, Golgi localization of tyrosylprotein sulfotransferase was sup- we characterized tyrosylprotein sulfotransferase from bovine ported by the observation that tyrosine sulfation of proteins in adrenal medullary Golgi fractions. The present results have intact cells was blocked by monensin and was in line with been reported in abstract form (14). previous observations that all tyrosine-sulfated proteins known so far are secretory. Glu,Ala,Tyr was found to have a very high affinity for tyrosylprotein sulfotransferase (apparent Kmi 300 METHODS nM), similar to that reported for certain tyrosylprotein kinases. Cell Cultures. Cells were grown and harvested as described While this may suggest some similarity between these enzymes, (8). Homogenates were prepared by adding 4 vol of Hepes the Golgi localization of tyrosylprotein sulfotransferase segre- solution (10 mM Hepes, pH 7.3/5 mM 2-mercaptoethanol) to gates tyrosine sulfation from the sites of tyrosine phosphoryl- 1 vol of cell pellets, followed by 10 strokes at 3000 rpm in a ation of proteins in the intact cell. If, however, tyrosylprotein glass/Teflon homogenizer. Homogenates were centrifuged sulfotransferase was allowed to react with cytoplasmic proteins for 1 hr at 165,000 x g and aliquots of the resulting pellets by using a nonionic detergent, tyrosine sulfation of tubulin was (total membrane fraction) were used in the standard sulfation observed. reaction described below. Subcellular Frationation. Subcellular fractionation of adult Previous reports from this laboratory have established that bovine adrenal medulla was done essentially as described by sulfation of proteins on tyrosine residues is a ubiquitous Trifaro and Duerr (15), except that 5 mM 2-mercaptoethanol posttranslational modification in animal cells (1-6). Various was used throughout. Briefly, the homogenate was centri- lines of evidence have indicated that tyrosine-sulfated pro- fuged at 800 x g for 10 min, and the resulting postnuclear teins belong mostly, if not exclusively, to one topological supernatant was centrifuged at 20,000 x g for 20 min. The class: the secretory proteins (see refs. 2-7 and references resulting pellet, the crude granule fraction (15), was then therein). One important approach to understanding the role of subfractionated by discontinuous sucrose density gradient tyrosine sulfation ofsecretory proteins is the characterization centrifugation to obtain the Golgi-enriched fraction (0.8/1.0 of the enzyme(s) catalyzing this protein modification. We M sucrose interface) (15). The supernatant obtained after the previously described an enzymatic activity in rat pheochro- centrifugation at 20,000 x g was centrifuged at 114,000 x g that transferred sulfate for 70 min to yield the soluble fraction and the crude mocytoma (PC12) cell membranes to microsomal fraction, which was resuspended in sucrose tyrosine residues of proteins, and we designated this enzyme solution (0.3 M sucrose/5 mM 2-mercaptoethanol). Aliquots tyrosylprotein sulfotransferase (8). This enzyme catalyzed ofeach subcellular fraction were stored at -20°C. Galactosyl sulfation of the same proteins that were tyrosine-sulfated by transferase activity in the various subcellular fractions was intact cells (8). The cosubstrate for sulfation was 3'- measured as described (16). phosphoadenosine 5'-phosphosulfate (PAPS), the universal Solubilization of Tyrosylprotein Sulfotransferase. Aliquots sulfate donor discovered by Lipmann and colleagues (for ofthe Golgi-enriched fraction were subjected to the following review, see ref. 9). three treatments. (i) The Golgi-enriched fraction (1 ml, In this study, we used the acidic random polymer contain- corresponding to 1.4 mg of protein) was diluted with 9 ml of ing tyrosine, (Glu62,Ala30,Tyr8), (referred to as Glu,Ala,Tyr), Hepes solution and then centrifuged at 165,000 x g for 60 as exogenous "protein" substrate to characterize tyrosyl- min. The supernatant was concentrated to 1 ml by vacuum protein sulfotransferase. The reasons for choosing Glu,Ala,Tyr dialysis against Hepes solution. The pellet was resuspended in 1 ml ofHepes solution. (ii) The Golgi-enriched fraction was The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" Abbreviation: PAPS, 3'-phosphoadenosine 5'-phosphosulfate. in accordance with 18 U.S.C. §1734 solely to indicate this fact. *To whom reprint requests should be addressed. 6143 Downloaded by guest on September 26, 2021 6144 Cell Biology: Lee and Huttner Proc. Natl. Acad. Sci. USA 82 (1985) treated as in i, except it was diluted with Hepes solution accessibility of PAPS to sulfotransferases, all reactions were containing 150 mM NaCl, and this suspension, prior to performed in the presence of nonionic detergent. In the centrifugation, was sonicated four times for 15 sec on ice at primary subfractions (Fig. 2, lanes A-D), most of the endog- maximum power, using a Branson B 15 Sonifier. (iii) The enous sulfation was found in the crude granule fraction [Fig. Golgi-enriched fraction was treated as in i, except that after 2, lane B(-)]. Subfractionation of the crude granule fraction centrifugation the pellet was resuspended in Hepes solution on a discontinuous sucrose density gradient (Fig. 2, lanes with 30% (wt/vol) glycerol/1% (wt/vol) Triton X-100, to a E-K) showed that most of the endogenous sulfation was protein concentration of 3 mg/ml, kept on ice for 1 hr, and observed in the fraction collected at the 0.8/1.0 M sucrose centrifuged at 165,000 x g for 60 min. The supernatant was interface [Fig. 2, lane G(-)]. This fraction is known to be collected and the pellet was resuspended in Hepes solution enriched in Golgi membrane vesicles, as determined by containing glycerol and Triton X-100 to the volume of the assaying for galactosyl transferase (ref. 15 and Fig. 2 Lower), supernatant. Aliquots (25 1.l) of the supernatant and pellet a trans-Golgi marker (19). A small amount of endogenous from treatments i-iii were then assayed for tyrosylprotein sulfation with a pattern similar to that of the Golgi-enriched sulfotransferase activity as described below. fraction was observed in the fraction collected at the 0.3/0.8 Tyrosylprotein Sulfotransferase Assay. Prior to the assay, M sucrose interface [Fig. 2, lane F(-)]. Approximately 30% the ethanol in [35S]PAPS (New England Nuclear; 0.5-3 of the total 35SO4 incorporated into proteins of the Golgi- Ci/mmol; 1 Ci = 37 GBq) was evaporated under a stream of enriched fraction [Fig. 2, lane G(-)] was recovered as nitrogen. The standard reaction mixture [see Fig. 2, lane tyrosine 35S4. The remaining 70% was alkali labile and was, G(+)] of 100 pl was composed of, in final concentrations, 10 therefore, presumably carbohydrate 35SO4. A different pat- mM Hepes, pH 7.0/5 mM MgCl2/5 mM MnCl2/5 mM tern of endogenous sulfation was observed in the fractions of 2-mercaptoethanol/1 mM NaF/20 1M [35S]PAPS/1% Triton the sucrose gradient enriched in chromaffin granules [Fig. 2, X-100/4 ,M Glu,Ala,Tyr (Sigma, P 3899; average size, 25 lanes J(-) and K(-)]. More than 97% of this incorporated kDa) and 70 ul of Golgi-enriched fraction (0.7-1.5 mg of 35 O4 appeared to be linked to carbohydrate residues. No protein per ml), which was added last to start the reaction. significant endogenous sulfation was observed in fractions The reaction mixture was incubated
Load more

Recommended publications
  • ECO-Ssls for Pahs
    Ecological Soil Screening Levels for Polycyclic Aromatic Hydrocarbons (PAHs) Interim Final OSWER Directive 9285.7-78 U.S. Environmental Protection Agency Office of Solid Waste and Emergency Response 1200 Pennsylvania Avenue, N.W. Washington, DC 20460 June 2007 This page intentionally left blank TABLE OF CONTENTS 1.0 INTRODUCTION .......................................................1 2.0 SUMMARY OF ECO-SSLs FOR PAHs......................................1 3.0 ECO-SSL FOR TERRESTRIAL PLANTS....................................4 5.0 ECO-SSL FOR AVIAN WILDLIFE.........................................8 6.0 ECO-SSL FOR MAMMALIAN WILDLIFE..................................8 6.1 Mammalian TRV ...................................................8 6.2 Estimation of Dose and Calculation of the Eco-SSL ........................9 7.0 REFERENCES .........................................................16 7.1 General PAH References ............................................16 7.2 References Used for Derivation of Plant and Soil Invertebrate Eco-SSLs ......17 7.3 References Rejected for Use in Derivation of Plant and Soil Invertebrate Eco-SSLs ...............................................................18 7.4 References Used in Derivation of Wildlife TRVs .........................25 7.5 References Rejected for Use in Derivation of Wildlife TRV ................28 i LIST OF TABLES Table 2.1 PAH Eco-SSLs (mg/kg dry weight in soil) ..............................4 Table 3.1 Plant Toxicity Data - PAHs ..........................................5 Table 4.1
    [Show full text]
  • Biosynthesis and Secretion of the Microbial Sulfated Peptide Raxx and Binding to the Rice XA21 Immune Receptor
    Biosynthesis and secretion of the microbial sulfated peptide RaxX and binding to the rice XA21 immune receptor Dee Dee Luua,b,1, Anna Joea,b,c,1, Yan Chend, Katarzyna Paryse, Ofir Bahara,b,2, Rory Pruitta,b,3, Leanne Jade G. Chand, Christopher J. Petzoldd, Kelsey Longa,b, Clifford Adamchaka,b, Valley Stewartf, Youssef Belkhadire, and Pamela C. Ronalda,b,c,4 aDepartment of Plant Pathology, University of California, Davis, CA 95616; bThe Genome Center, University of California, Davis, CA 95616; cFeedstocks Division, Joint Bioenergy Institute, Emeryville, CA 94608; dTechnology Division, Joint Bioenergy Institute, Emeryville, CA 94608; eGregor Mendel Institute, Austrian Academy of Sciences, 1030 Vienna, Austria; and fDepartment of Microbiology & Molecular Genetics, University of California, Davis, CA 95616 Edited by Jonathan D. G. Jones, The Sainsbury Laboratory, Norwich, United Kingdom, and approved March 7, 2019 (received for review October 24, 2018) The rice immune receptor XA21 is activated by the sulfated micro- to Xoo strains lacking RaxX (7–10). Xoo strains lacking RaxX are bial peptide required for activation of XA21-mediated immunity X also compromised in virulence of rice plants lacking XA21 (10). (RaxX) produced by Xanthomonas oryzae pv. oryzae (Xoo). Muta- These results suggest a role for RaxX in bacterial virulence. tional studies and targeted proteomics revealed that the RaxX pre- Tyrosine sulfated RaxX16, a 16-residue synthetic peptide de- cursor peptide (proRaxX) is processed and secreted by the protease/ rived from residues 40–55 of the 60-residue RaxX precursor transporter RaxB, the function of which can be partially fulfilled by peptide (proRaxX), is the shortest characterized immunogenic a noncognate peptidase-containing transporter component B derivative of RaxX (10).
    [Show full text]
  • NDA/BLA Multi-Disciplinary Review and Evaluation
    NDA/BLA Multi-disciplinary Review and Evaluation NDA 214154 Nextstellis (drospirenone and estetrol tablets) NDA/BLA Multi-Disciplinary Review and Evaluation Application Type NDA Application Number(s) NDA 214154 (IND 110682) Priority or Standard Standard Submit Date(s) April 15, 2020 Received Date(s) April 15, 2020 PDUFA Goal Date April 15, 2021 Division/Office Division of Urology, Obstetrics, and Gynecology (DUOG) / Office of Rare Diseases, Pediatrics, Urologic and Reproductive Medicine (ORPURM) Review Completion Date April 15, 2021 Established/Proper Name drospirenone and estetrol tablets (Proposed) Trade Name Nextstellis Pharmacologic Class Combination hormonal contraceptive Applicant Mayne Pharma LLC Dosage form Tablet Applicant proposed Dosing x Take one tablet by mouth at the same time every day. Regimen x Take tablets in the order directed on the blister pack. Applicant Proposed For use by females of reproductive potential to prevent Indication(s)/Population(s) pregnancy Recommendation on Approval Regulatory Action Recommended For use by females of reproductive potential to prevent Indication(s)/Population(s) pregnancy (if applicable) Recommended Dosing x Take one pink tablet (drospirenone 3 mg, estetrol Regimen anhydrous 14.2 mg) by mouth at the same time every day for 24 days x Take one white inert tablet (placebo) by mouth at the same time every day for 4 days following the pink tablets x Take tablets in the order directed on the blister pack 1 Reference ID: 4778993 NDA/BLA Multi-disciplinary Review and Evaluation NDA 214154 Nextstellis (drospirenone and estetrol tablets) Table of Contents Table of Tables .................................................................................................................... 5 Table of Figures ................................................................................................................... 7 Reviewers of Multi-Disciplinary Review and Evaluation ...................................................
    [Show full text]
  • Biotransformation: Basic Concepts (1)
    Chapter 5 Absorption, Distribution, Metabolism, and Elimination of Toxics Biotransformation: Basic Concepts (1) • Renal excretion of chemicals Biotransformation: Basic Concepts (2) • Biological basis for xenobiotic metabolism: – To convert lipid-soluble, non-polar, non-excretable forms of chemicals to water-soluble, polar forms that are excretable in bile and urine. – The transformation process may take place as a result of the interaction of the toxic substance with enzymes found primarily in the cell endoplasmic reticulum, cytoplasm, and mitochondria. – The liver is the primary organ where biotransformation occurs. Biotransformation: Basic Concepts (3) Biotransformation: Basic Concepts (4) • Interaction with these enzymes may change the toxicant to either a less or a more toxic form. • Generally, biotransformation occurs in two phases. – Phase I involves catabolic reactions that break down the toxicant into various components. • Catabolic reactions include oxidation, reduction, and hydrolysis. – Oxidation occurs when a molecule combines with oxygen, loses hydrogen, or loses one or more electrons. – Reduction occurs when a molecule combines with hydrogen, loses oxygen, or gains one or more electrons. – Hydrolysis is the process in which a chemical compound is split into smaller molecules by reacting with water. • In most cases these reactions make the chemical less toxic, more water soluble, and easier to excrete. Biotransformation: Basic Concepts (5) – Phase II reactions involves the binding of molecules to either the original toxic molecule or the toxic molecule metabolite derived from the Phase I reactions. The final product is usually water soluble and, therefore, easier to excrete from the body. • Phase II reactions include glucuronidation, sulfation, acetylation, methylation, conjugation with glutathione, and conjugation with amino acids (such as glycine, taurine, and glutamic acid).
    [Show full text]
  • A Novel Method for Site-Determination of Tyrosine O- Sulfation in Peptides and Proteins
    A novel method for site-determination of tyrosine O- sulfation in peptides and proteins Yonghao Yu Genome Center, Depts of Chemistry and Molecular Cell Biology, University of California, Davis, CA, 95616 and Dept of Chemistry, University of California, Berkeley, CA, 94720, current address: Dept of Cell Biology, Harvard Medical School, Boston, MA 02115 Adam J. Hoffhines Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104 Kevin L. Moore Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104 Julie A. Leary Genome Center, Departments of Chemistry and Molecular Cell Biology, University of California, Davis, CA, 95616 Method Article Keywords: tyrosine sulfation, mass spectrometry, post-translational modication, protein-protein interaction, extracelluar matrix, proteomics Posted Date: September 20th, 2007 DOI: https://doi.org/10.1038/nprot.2007.388 License: This work is licensed under a Creative Commons Attribution 4.0 International License. Read Full License Page 1/12 Abstract Introduction Tyrosine O-sulfation is one of many posttranslational modications described in nature that can impart critical functional properties to proteins that are independent of the genes encoding them1, 2. It plays a key role in regulating protein-protein interactions in the extracellular space3 and is catalyzed by two closely related Golgi enzymes called tyrosylprotein sulfotransferases \(TPST-1 and TPST–2)4, 5. In this protocol, we describe a novel subtractive strategy to determine the sites of tyrosine sulfation in peptides and proteins6. Hydroxyl groups on unsulfated tyrosines are stoichiometrically acetylated by a one-step reaction using sulfosuccinimidyl acetate in the presence of imidazole at pH 7.0.
    [Show full text]
  • Tyrosine Sulfation: a Modulator of Extracellular Protein–Protein Interactions Provided by Elsevier - Publisher Connector John W Kehoe1 and Carolyn R Bertozzi1,2
    cm7309.qxd 03/10/2000 11:46 Page R57 CORE Minireview R57 brought to you by Elsevier - Publisher Connector Tyrosine sulfation: a modulator of extracellular protein–protein interactions provided by John W Kehoe1 and Carolyn R Bertozzi1,2 Tyrosine sulfation is a post-translational modification of P-selectin on activated endothelial cells with the many secreted and membrane-bound proteins. Its P-selectin glycoprotein ligand (PSGL)-1 on cognate biological roles have been unclear. Recent work has leukocytes [3]. PSGL-1 was identified at the molecular implicated tyrosine sulfate as a determinant of level in 1993, using expression cloning [4]. Glycosylation protein–protein interactions involved in leukocyte of PSGL-1 was known to be a major determinant of adhesion, hemostasis and chemokine signaling. P-selectin binding, but was not sufficient to explain the binding avidities observed in vivo. Addresses: Departments of 1Molecular and Cell Biology and 2 Chemistry, University of California, Berkeley, CA 94720, USA. Further investigations revealed that tyrosine sulfation near Correspondence: Carolyn R Bertozzi the amino terminus of PSGL-1 plays an essential role in E-mail: [email protected] P-selectin binding (shown schematically in Figure 1b). Three papers published within one month of each other in Chemistry & Biology 2000, 7:R57–R61 1995 provided the first evidence of this phenomenon 1074-5521/00/$ – see front matter [5–7]. Progressive deletions of the PSGL-1 amino termi- © 2000 Elsevier Science Ltd. All rights reserved. nus by Pouyani and Seed [5] revealed that a 20-residue peptide was required for high-affinity binding. Further- The study of living systems has been transformed by an more, mutagenesis within this sequence showed that the explosion of genetic information.
    [Show full text]
  • NST110: Advanced Toxicology Lecture 4: Phase I Metabolism
    Absorption, Distribution, Metabolism and Excretion (ADME): NST110: Advanced Toxicology Lecture 4: Phase I Metabolism NST110, Toxicology Department of Nutritional Sciences and Toxicology University of California, Berkeley Biotransformation The elimination of xenobiotics often depends on their conversion to water-soluble chemicals through biotransformation, catalyzed by multiple enzymes primarily in the liver with contributions from other tissues. Biotransformation changes the properties of a xenobiotic usually from a lipophilic form (that favors absorption) to a hydrophilic form (favoring excretion in the urine or bile). The main evolutionary goal of biotransformation is to increase the rate of excretion of xenobiotics or drugs. Biotransformation can detoxify or bioactivate xenobiotics to more toxic forms that can cause tumorigenicity or other toxicity. Phase I and Phase II Biotransformation Reactions catalyzed by xenobiotic biotransforming enzymes are generally divided into two groups: Phase I and phase II. 1. Phase I reactions involve hydrolysis, reduction and oxidation, exposing or introducing a functional group (-OH, -NH2, -SH or –COOH) to increase reactivity and slightly increase hydrophilicity. O R1 - O S O sulfation O R2 OH Phase II Phase I R1 R2 R1 R2 - hydroxylation COO R1 O O glucuronidation OH R2 HO excretion OH O COO- HN H -NH2 R R1 R1 S N 1 Phase I Phase II COO O O oxidation glutathione R2 OH R2 R2 conjugation 2. Phase II reactions include glucuronidation, sulfation, acetylation, methylation, conjugation with glutathione, and conjugation with amino acids (glycine, taurine and glutamic acid) that strongly increase hydrophilicity. Phase I and II Biotransformation • With the exception of lipid storage sites and the MDR transporter system, organisms have little anatomical defense against lipid soluble toxins.
    [Show full text]
  • 6. References
    332 IARC MONOGRAPHS VOLUME 91 6. References Abramov, Y., Borik, S., Yahalom, C., Fatum, M., Avgil, G., Brzezinski, A. & Banin, E. (2004) The effect of hormone therapy on the risk for age-related maculopathy in postmenopausal women. Menopause, 11, 62–68 Adams, M.R., Register, T.C., Golden, D.L., Wagner, JD. & Williams, J.K. (1997) Medroxyproges- terone acetate antagonizes inhibitory effects of conjugated equine estrogens on coronary artery atherosclerosis. Arterioscler. Thromb. vasc. Biol., 17, 217–221 Adjei, A.A. & Weinshilboum, R.M. (2002) Catecholestrogen sulfation: Possible role in carcino- genesis. Biochem. biophys. Res. Commun., 292, 402–408 Adjei, A.A., Thomae, B.A., Prondzinski, J.L., Eckloff, B.W., Wieben, E.D. & Weinshilboum, R.M. (2003) Human estrogen sulfotransferase (SULT1E1) pharmacogenomics: Gene resequencing and functional genomics. Br. J. Pharmacol., 139, 1373–1382 Ahmad, M.E., Shadab, G.G.H.A., Hoda, A. & Afzal, M. (2000) Genotoxic effects of estradiol-17β on human lymphocyte chromosomes. Mutat. Res., 466, 109–115 COMBINED ESTROGEN−PROTESTOGEN MENOPAUSAL THERAPY 333 Ahmad, M.E., Shadab, G.G.H.A., Azfer, M.A. & Afzal, M. (2001) Evaluation of genotoxic poten- tial of synthetic progestins-norethindrone and norgestrel in human lymphocytes in vitro. Mutat. Res., 494, 13–20 Aitken, J.M., Hart, D.M. & Lindsay, R. (1973) Oestrogen replacement therapy for prevention of osteoporosis after oophorectomy. Br. med. J., 3, 515–518 Al-Azzawi, F., Wahab, M., Thompson, J., Whitehead, M. & Thompson, W. (1999) Acceptability and patterns of uterine bleeding in sequential trimegestone-based hormone replacement therapy: A dose-ranging study. Hum. Reprod., 14, 636–641 Albert, C., Vallée, M., Beaudry, G., Belanger, A.
    [Show full text]
  • Estrogen Metabolism: Are We Assessing It Properly? Filomena Trindade, MD, MPH
    Estrogen Metabolism: Are We Assessing It Properly? Filomena Trindade, MD, MPH The views and opinions expressed herein are solely those of the presenter and do not necessarily represent those of Genova Diagnostics. Thus, Genova Diagnostics does not accept liability for consequences of any actions taken on the basis of the information provided. Michael Chapman, ND Medical Education Specialist - Asheville Filomena Trindade, MD, MPH Technical Issues & Clinical Questions Please type any technical issue or clinical question into either the “Chat” or “Questions” boxes, making sure to send them to “Organizer” at any time during the webinar. We will be compiling your clinical questions and answering as many as we can the final 15 minutes of the webinar. DISCLAIMER: Please note that any and all emails provided may be used for follow up correspondence and/or for further communication. Estrogen Metabolism: Are We Assessing It Properly? Filomena Trindade, MD, MPH The views and opinions expressed herein are solely those of the presenter and do not necessarily represent those of Genova Diagnostics. Thus, Genova Diagnostics does not accept liability for consequences of any actions taken on the basis of the information provided. Objectives • Understand the importance of estrogen metabolism with respect to cancer risk • Be able to devise a treatment plan for a pt with an unfavorable estrogen metabolism profile • Gain a basic understanding of the importance of methylation in estrogen metabolism, health promotion and cancer prevention • Learn to apply these
    [Show full text]
  • Intratumoral Estrogen Sulfotransferase Induction Contributes to the Anti-Breast Cancer Effects of the Dithiocarbamate Derivative TM208
    npg Acta Pharmacologica Sinica (2015) 36: 1246–1255 © 2015 CPS and SIMM All rights reserved 1671-4083/15 www.nature.com/aps Original Article Intratumoral estrogen sulfotransferase induction contributes to the anti-breast cancer effects of the dithiocarbamate derivative TM208 Xi-wei JI 1, 2, Guang-ping CHEN4, Yan SONG5, Ming HUA1, Li-jie WANG1, Liang LI1, Yin YUAN1, Si-yuan WANG1, Tian-yan ZHOU1, 3, *, Wei LU1, 3 1Department of Pharmaceutics, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China; 2Institute of Clinical Pharmacology, Peking University First Hospital, Beijing 100191, China; 3State Key Laboratory of Natural and Biomimetic Drugs, Peking University, Beijing 100191, China; 4Department of Physiological Sciences, Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, OK 74078, USA; 5Department of Molecular and Cellular Pharmacology, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China Aim: Sulfotransferase-catalyzed sulfation is the most important pathway for inactivating estrogens. Thus, activation of estrogen sulfotransferase (EST) may be an alternative approach for the treatment of estrogen-dependent breast cancer. In this study we investigated the involvement of EST in anti-breast cancer effects of the dithiocarbamate derivative TM208 in vitro and in vivo. Methods: The viability of human breast cancer MCF-7 cells was determined using a SBB assay. Nude mice bearing MCF-7 cells were orally administered TM208 (50 and 150 mg·kg-1·d-1) for 18 days. The xenograft tumors and uteri were collected. The mRNA expression of EST was examined with real-time PCR. EST protein was detected with Western blot, ELISA or immunohistochemical staining assays. A radioactive assay was used to measure the EST activity.
    [Show full text]
  • Nutritional Flavonoids Impact on Nuclear and Extranuclear Estrogen Receptor Activities
    Genes & Nutrition Vol. 1, No. 3/4, pp. 161-176, 2006 ISSN 1555-8932 print, Copyright © 2006 by New Century Health Publishers, LLC www.newcenturyhealthpublishers.com All rights of reproduction in any form reserved NUTRITIONAL FLAVONOIDS IMPACT ON NUCLEAR AND EXTRANUCLEAR ESTROGEN RECEPTOR ACTIVITIES Paola Galluzzo and Maria Marino Department of Biology, University “Roma Tre”, Viale G. Marconi 446, I-00146 Roma, Italy [Received March 14, 2006; Accepted April 30, 2006] ABSTRAABSTRAABSTRACTCTCT: Flavonoids are a large group of non-nutrient important are the plant secondary metabolites flavonoids. They compounds naturally produced from plants as part of their are present in all terrestrial vascular plants whereas, in mammals, defence mechanisms against stresses of different origins. They they occur only through dietary intake (Birt et al., 2001; Dixon, emerged from being considered an agricultural oddity only after 2004). More than 4,000 different flavonoids have been it was observed that these compounds possess a potential protective described and categorized into 6 subclasses as a function of function against several human degenerative diseases. This has the type of heterocycle involved: flavonols, flavones, flavanols, led to recommending the consumption of food containing high flavanonols, flavanones, and isoflavones (Table 1) (Birt et al., concentrations of flavonoids, which at present, especially as soy 2001; Manach et al., 2004). isoflavones, are even available as over-the-counter nutraceuticals. The increased use of flavonoids has occurred even though their mechanisms are not completely understood, in particular those TABLE 11:1TABLE Chemical structures of certain commonly occurring plant involving the flavonoid impact on estrogen signals. In fact, most flavonoid aglycones of the human health protective effects of flavonoids are described either as estrogen-mimetic, or as anti-estrogenic, while others do not involve estrogen signaling at all.
    [Show full text]
  • Sequential Tyrosine Sulfation of CXCR4 by Tyrosylprotein Sulfotransferases† Christoph Seibert,*,‡ Christopher T
    Biochemistry XXXX, xxx, 000–000 A Sequential Tyrosine Sulfation of CXCR4 by Tyrosylprotein Sulfotransferases† Christoph Seibert,*,‡ Christopher T. Veldkamp,§ Francis C. Peterson,§ Brian T. Chait,| Brian F. Volkman,§ and Thomas P. Sakmar‡ Laboratory of Molecular Biology and Biochemistry and Laboratory for Mass Spectrometry and Gaseous Ion Chemistry, The Rockefeller UniVersity, New York, New York 10065, and Department of Biochemistry, Medical College of Wisconsin, Milwaukee, Wisconsin 53226 ReceiVed May 21, 2008; ReVised Manuscript ReceiVed August 27, 2008 ABSTRACT: CXC-chemokine receptor 4 (CXCR4) is a G protein-coupled receptor for stromal cell-derived factor-1 (SDF-1/CXCL12). SDF-1-induced CXCR4 signaling is indispensable for embryonic development and crucial for immune cell homing and has been implicated in metastasis of numerous types of cancer. CXCR4 also serves as the major coreceptor for cellular entry of T-cell line-tropic (X4) HIV-1 strains. Tyrosine residues in the N-terminal tail of CXCR4, which are post-translationally sulfated, are implicated in the high-affinity binding of SDF-1 to CXCR4. However, the specific roles of three potential tyrosine sulfation sites are not well understood. We investigated the pattern and sequence of CXCR4 sulfation by using recombinant human tyrosylprotein sulfotransferases TPST-1 and TPST-2 to modify a peptide that corresponds to amino acids 1-38 of the receptor (CXCR4 1-38). We analyzed the reaction products with a combination of reversed-phase HPLC, proteolytic cleavage, and mass spectrometry. We found that CXCR4 1-38 is sulfated efficiently by both TPST enzymes, leading to a final product with three sulfotyrosine residues. Sulfates were added stepwise to the peptide, producing specific intermediates with one or two sulfotyrosines.
    [Show full text]