DOCSLIB.ORG

Arabidopsis and Chlamydomonas Phosphoribulokinase Crystal Structures Complete the Redox Structural Proteome of the Calvin-Benson

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Arabidopsis and Chlamydomonas Phosphoribulokinase Crystal Structures Complete the Redox Structural Proteome of the Calvin-Benson bioRxiv preprint doi: https://doi.org/10.1101/422709; this version posted February 18, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 1 Classification: Biological Sciences: Plant Biology 2 3 Arabidopsis and Chlamydomonas phosphoribulokinase crystal structures complete the redox 4 structural proteome of the Calvin-Benson cycle 5 6 Short title: Insight into chloroplast PRK structure 7 8 Libero Gurrieria, Alessandra Del Giudiceb, Nicola Demitric, Giuseppe Falinid, Nicolae Viorel Pavelb, 9 Mirko Zaffagninia, Maurizio Polentaruttic, Pierre Crozete, Christophe H. Marchande, Julien Henrie, 10 Paolo Trosta, Stéphane D. Lemairee, Francesca Sparlaa,1, Simona Fermanid,1 11 12 aDepartment of Pharmacy and Biotechnology – FaBiT, University of Bologna, 40126 Bologna, Italy 13 bDepartment of Chemistry, University of Rome ‘Sapienza’, 00185 Rome, Italy 14 cElettra - Sincrotone Trieste, 34149, Basovizza - Trieste, Italy 15 dDepartment of Chemistry ‘G. Ciamician’, University of Bologna, 40126 Bologna, Italy 16 eInstitut de Biologie Physico-Chimique, UMR8226, CNRS, Sorbonne Université, 75005 Paris, 17 France 18 19 20 1To whom correspondence should be addressed: 21 Simona Fermani: 22 Address: Department of Chemistry ‘G. Ciamician,’ University of Bologna, Via Selmi 2 - 23 40126 Bologna, Italy 24 Phone: +39 051 2099475 25 Email address: [email protected] 26 Francesca Sparla: 1 bioRxiv preprint doi: https://doi.org/10.1101/422709; this version posted February 18, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 27 Address: Department of Pharmacy and Biotechnology – FaBiT, University of Bologna, Via 28 Irnerio 42 - 40126 Bologna, Italy 29 Phone: +39 051 2091281 30 Email address: [email protected] 2 bioRxiv preprint doi: https://doi.org/10.1101/422709; this version posted February 18, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 31 Abstract 32 In land plants and algae, the Calvin-Benson (CB) cycle takes place in the chloroplast, a specialized 33 organelle in which photosynthesis occurs. Thioredoxins (TRXs) are small ubiquitous proteins, 34 known to harmonize the two stages of photosynthesis through a thiol-based mechanism. Among the 35 11 enzymes of the CB cycle, the TRX target phosphoribulokinase (PRK) has yet to be characterized 36 at the atomic scale. To accomplish this goal, we determined the crystal structures of PRK from two 37 model species: the green alga Chlamydomonas reinhardtii (CrPRK) and the land plant Arabidopsis 38 thaliana (AtPRK). PRK is an elongated homodimer characterized by a large central β-sheet of 18 39 strands, extending between two catalytic sites positioned at its edges. The electrostatic surface 40 potential of the catalytic cavity has both a positive region suitable for binding the phosphate groups 41 of substrates and an exposed negative region to attract positively charged TRX-f. In the catalytic 42 cavity, the regulatory cysteines are 13 Å apart and connected by a flexible region exclusive to 43 photosynthetic eukaryotes—the clamp loop—which is believed to be essential for oxidation- 44 induced structural rearrangements. Structural comparisons with prokaryotic and evolutionarily older 45 PRKs revealed that both AtPRK and CrPRK have a strongly reduced dimer interface and increased 46 number of random coiled regions, suggesting that a general loss in structural rigidity correlates with 47 gains in TRX sensitivity during the molecular evolution of PRKs in eukaryotes. 48 49 Keywords: 3D structure; Calvin-Benson cycle; phosphoribulokinase; redox-regulation; structural 50 flexibility; thioredoxin 51 52 Significance Statement 53 In chloroplasts, five enzymes of the Calvin-Benson (CB) cycle are regulated by thioredoxins 54 (TRXs). These enzymes have all been structurally characterized with the notable exception of 55 phosphoribulokinase (PRK). Here, we determined the crystal structure of chloroplast PRK from two 56 model photosynthetic organisms. Regulatory cysteines appear distant from each other and are 3 bioRxiv preprint doi: https://doi.org/10.1101/422709; this version posted February 18, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 57 linked by a long loop that is present only in plant-type PRKs and allows disulfide bond formation 58 and subsequent conformational rearrangements. Structural comparisons with ancient PRKs indicate 59 that the presence of flexible regions close to regulatory cysteines is a unique feature that is shared 60 by TRX-dependent CB cycle enzymes, suggesting that the evolution of the PRK structure has 61 resulted in a global increase in protein flexibility for photosynthetic eukaryotes. 62 63 INTRODUCTION 64 Photosynthetic CO2 fixation supports life on earth and is a fundamental source of food, fuel, and 65 chemicals for human society. In the vast majority of photosynthetic organisms, carbon fixation 66 occurs via the Calvin-Benson (CB) cycle (Fig. S1), a pathway that has been extensively studied at 67 the physiological, biochemical, and structural levels. The CB cycle consists of 13 distinct reactions 68 catalyzed by 11 enzymes (1) that are differentially regulated to coordinate the two stages of 69 photosynthesis—electron transport and carbon fixation—and to couple them with the continuous 70 changes in environmental light. 71 Thioredoxins (TRXs) are small thiol oxidoreductase enzymes that are widely distributed among the 72 kingdoms of life. They reduce disulfide bonds in target proteins, controlling the redox state and 73 modulating the activity of target enzymes (2). Compared to animals that usually possess 2–3 74 different TRXs, the plant thioredoxin system is much more complex. Looking only at the 75 chloroplast, five different classes of TRXs are known (i.e., TRX-f, -m, -x, -y, and -z) with several 76 isoforms composing many of these classes (2). Despite this diversity, TRXs are ancient and 77 strongly evolutionarily conserved, consisting of a central core of a five-stranded β-sheet surrounded 78 by four α-helices and an active site that protrudes from the protein’s surface (3). 79 In TRX-dependent regulation, a small fraction of photosynthetic-reducing equivalents are 80 transferred from photosystem I to several chloroplast TRXs, which in turn reduce key enzymes of 81 the CB cycle, regulating their activity (1). Among the five classes of chloroplast TRXs, class f 82 displays a high specificity towards CB cycle enzymes. In both plants and algae, the activities of 4 bioRxiv preprint doi: https://doi.org/10.1101/422709; this version posted February 18, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 83 fructose-1,6-bisphosphatase (FBPase) (4, 5), sedoheptulose-1,7-bisphosphatase (SBPase) (5), PRK 84 (6), and the AB isoform of NADP-glyceraldehyde 3-phosphate dehydrogenase (AB-GAPDH) (7) in 85 plants and of phosphoglycerate kinase (PGK) (8) in Chlamydomonas are directly linked to light by 86 a thiol-based mechanism driven by TRX (Fig. S1). In addition, PRK forms a regulatory complex 87 with the redox-sensitive CP12 and the homotetrameric NADP-GAPDH (A4-GAPDH) (9-11). The 88 formation of this ternary complex occurs in both plants and algae, and causes an almost complete 89 inhibition of PRK activity (12). 90 In contrast to the canonical WCGPC active site of TRXs, no thioredoxin-recognition signature is 91 found in these enzymes. A structural comparison of SBPase and FBPase highlights how evolution 92 has adopted different strategies for modeling protein structure to achieve the same TRX-dependent 93 redox control (5). Of the three enzymes regulated by TRX whose 3D structures are available, a pair 94 of redox-sensitive cysteines was found to be located in completely different regions. In AB- 95 GAPDH, these cysteines are located in a C-terminal extension (CTE) (7); in SBPase, they are 96 present at the dimer interface (5); and in FBPase, they are solvent exposed and distant from the 97 sugar-binding site (4, 5). 98 To complete the structural redox proteome of the CB cycle enzymes and identify the common 99 feature behind the acquisition of TRX-mediated redox control, the crystal structures of PRK from 100 two model species—the green alga Chlamydomonas reinhardtii and the land plant Arabidopsis 101 thaliana—were determined and compared to those of other TRX-controlled enzymes. Our results 102 strongly show that the acquisition of a TRX-targeted motif positively correlates with the acquisition 103 of flexibility in the target proteins, consistent with the observation that the entropic contribution 104 obtained from the reduction of the oxidized target enzyme is the main driving force of the redox 105 control mediated by TRXs (13). 106 107 RESULTS AND DISCUSSION 108 Three-dimensional structures of CrPRK and AtPRK in solution and in crystal form 5 bioRxiv preprint doi: https://doi.org/10.1101/422709; this version posted February 18, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.
Load more

Recommended publications
  • Allosteric Regulation in Drug Design
    Mini Review Curr Trends Biomedical Eng & Biosci Volume 4 Issue 1 - May 2017 Copyright © All rights are reserved by Ashfaq Ur Rehman DOI: 10.19080/CTBEB.2017.04.5555630 Allosteric regulation in drug design Ashfaq Ur Rehman1,2*, Shah Saud3, Nasir Ahmad4, Abdul Wadood2 and R Hamid5 1State Key Laboratory of Microbial Metabolism, Department of Bioinformatics and Biostatistics, China 2Department of Biochemistry, Abdul Wali Khan University Mardan, Pakistan 3Laboratory of Analytical Biochemistry and Bio separation, Shanghai Jiao Tong University, China 4Department of Chemistry, Islama College University Peshawar, Pakistan 5Department of Bioinformatics, Muhammad Ali Jinnah University Islamabad, Pakistan Submission: May 02, 2017; Published: May 23, 2017 *Corresponding author: Ashfaq Ur Rehman, State Key Laboratory of Microbial Metabolism, Department of Bioinformatics and Biostatistics, Shanghai Jiao Tong University, 800 Dongchuan Road, Shanghai 200240, China, Tel: ; Fax: 86-21-34204348; Email: Abstract mechanism, which are initiated through attachment of ligand or inhibitors with the protein or enzymes other than active (orthosteric) sites. ThisProtein mini review and enzymes involved play mechanism, significant types roles and in importancebiological processes of allosteric of all regulations living organisms; in drug theirdesign functions process. are regulated through allosteric Keywords: Allosteric, Activator: Drug design Introduction and ultimately cause disease. While various biological processes expressed the control at different points in life time of protein function is pivotal. As all the cell processes are under carful For the survival of all organisms the significance of protein included regulation of gene expression, translation into protein control and if not properly controls this leads to the abnormality through control of activity and at last degradation of protein [1].
    [Show full text]
  • On the Active Site Thiol of Y-Glutamylcysteine Synthetase
    Proc. Natl. Acad. Sci. USA Vol. 85, pp. 2464-2468, April 1988 Biochemistry On the active site thiol of y-glutamylcysteine synthetase: Relationships to catalysis, inhibition, and regulation (glutathione/cystamine/Escherichia coli/kidney/enzyme inactivation) CHIN-SHIou HUANG, WILLIAM R. MOORE, AND ALTON MEISTER Cornell University Medical College, Department of Biochemistry, 1300 York Avenue, New York, NY 10021 Contributed by Alton Meister, December 4, 1987 ABSTRACT y-Glutamylcysteine synthetase (glutamate- dithiothreitol, suggesting that cystamine forms a mixed cysteine ligase; EC 6.3.2.2) was isolated from an Escherichia disulfide between cysteamine and an enzyme thiol (15). coli strain enriched in the gene for this enzyme by recombinant Inactivation of the enzyme by the L- and D-isomers of DNA techniques. The purified enzyme has a specific activity of 3-amino-1-chloro-2-pentanone, as well as that by cystamine, 1860 units/mg and a molecular weight of 56,000. Comparison is prevented by L-glutamate (14). Treatment of the enzyme of the E. coli enzyme with the well-characterized rat kidney with cystamine prevents its interaction with the sulfoxi- enzyme showed that these enzymes have similar catalytic prop- mines. Titration of the enzyme with 5,5'-dithiobis(2- erties (apparent Km values, substrate specificities, turnover nitrobenzoate) reveals that the enzyme has a single exposed numbers). Both enzymes are feedback-inhibited by glutathione thiol that reacts with this reagent without affecting activity but not by y-glutamyl-a-aminobutyrylglycine; the data indicate (16). 5,5'-Dithiobis(2-nitrobenzoate) does not interact with that glutathione binds not only at the glutamate binding site but the thiol that reacts with cystamine.
    [Show full text]
  • Molecular Markers of Serine Protease Evolution
    The EMBO Journal Vol. 20 No. 12 pp. 3036±3045, 2001 Molecular markers of serine protease evolution Maxwell M.Krem and Enrico Di Cera1 ment and specialization of the catalytic architecture should correspond to signi®cant evolutionary transitions in the Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, Box 8231, St Louis, history of protease clans. Evolutionary markers encoun- MO 63110-1093, USA tered in the sequences contributing to the catalytic apparatus would thus give an account of the history of 1Corresponding author e-mail: [email protected] an enzyme family or clan and provide for comparative analysis with other families and clans. Therefore, the use The evolutionary history of serine proteases can be of sequence markers associated with active site structure accounted for by highly conserved amino acids that generates a model for protease evolution with broad form crucial structural and chemical elements of applicability and potential for extension to other classes of the catalytic apparatus. These residues display non- enzymes. random dichotomies in either amino acid choice or The ®rst report of a sequence marker associated with serine codon usage and serve as discrete markers for active site chemistry was the observation that both AGY tracking changes in the active site environment and and TCN codons were used to encode active site serines in supporting structures. These markers categorize a variety of enzyme families (Brenner, 1988). Since serine proteases of the chymotrypsin-like, subtilisin- AGY®TCN interconversion is an uncommon event, it like and a/b-hydrolase fold clans according to phylo- was reasoned that enzymes within the same family genetic lineages, and indicate the relative ages and utilizing different active site codons belonged to different order of appearance of those lineages.
    [Show full text]
  • Crystallography Captures Catalytic Steps in Human Methionine Adenosyltransferase Enzymes
    Crystallography captures catalytic steps in human methionine adenosyltransferase enzymes Ben Murraya,b, Svetlana V. Antonyuka, Alberto Marinab, Shelly C. Luc, Jose M. Matod, S. Samar Hasnaina,1, and Adriana L. Rojasb,1 aMolecular Biophysics Group, Institute of Integrative Biology, Faculty of Health and Life Sciences, University of Liverpool, Liverpool L69 7ZX, England; bStructural Biology Unit, Center for Cooperative Research in Biosciences, 48160 Derio, Spain; cDivision of Gastroenterology, Cedars-Sinai Medical Center, Los Angeles, CA 90048; and dCIC bioGUNE, CIBERehd, Parque Tecnologico Bizkaia, 801A-1.48160 Derio, Spain Edited by Gregory A. Petsko, Weill Cornell Medical College, New York, NY, and approved January 8, 2016 (received for review June 4, 2015) The principal methyl donor of the cell, S-adenosylmethionine catalytic mechanism (13, 14) in which the reaction is initiated (SAMe), is produced by the highly conserved family of methionine through a nucleophilic attack by the sulfur atom of methionine adenosyltranferases (MATs) via an ATP-driven process. These en- on the C5′ atom of ATP, which produces the intermediate tri- zymes play an important role in the preservation of life, and their polyphosphate (PPPi). Hydrolysis of the PPPi into pyrophos- dysregulation has been tightly linked to liver and colon cancers. We phate (PPi) and orthophosphate (Pi) then occurs (Fig. S1). present crystal structures of human MATα2 containing various bound A common feature of MAT enzymes is a gating loop that α – ligands, providing a “structural movie” of the catalytic steps. High- to flanks the active site (in human MAT 2 residues 113 131), atomic-resolution structures reveal the structural elements of the which has been postulated to act in a dynamic way to allow ac- enzyme involved in utilization of the substrates methionine and cess to the active site.
    [Show full text]
  • Enzyme Regulation What Factors Influence Enzymatic Activity?
    12/3/13 Enzyme Regulation What Factors Influence Enzymatic Activity? Principle means of regulating enzyme activity • Reversible, non-covalent (allosteric and simple- MM) – typically small molecules • Reversible, covalent • Protein-Protein interactions • Zymogen activation • Protein expression and degradation • Availability (both of enzyme and substrate) Reversible Noncovalent: Reversible Noncovalent Allosteric Simple activation and inhibition by small molecules – Action at "another site" substrate, natural regulators of enzymes Enzymes situated at key steps in metabolic pathways are modulated by allosteric effectors MM kinetics Km, Vmax – competitive, non These effectors are usually produced elsewhere in competitive… the pathway Effectors may be feed-forward activators or Substrate inhibition or activation feedback inhibitors Kinetics are sigmoid ("S-shaped") The availability of substrates and cofactors usually determines how fast the reaction goes As product accumulates, the apparent rate of the enzymatic reaction will decrease General Features of Allosteric Regulation Allosteric activation/inhibition In most metabolic pathways there is at least 1 key enzyme Usually these pacemaker enzymes are in the first committed step in the pathway. Many times regulated by both feed forward and feedback mechanisms A B C D E Sigmoid v versus [S] plot. The dotted line represents the hyperbolic plot characteristic of normal Michaelis=Menten kinetics. 1 12/3/13 Allosteric activation/inhibition Reversible Covalent In most metabolic pathways there is at
    [Show full text]
  • Understanding Drug-Drug Interactions Due to Mechanism-Based Inhibition in Clinical Practice
    pharmaceutics Review Mechanisms of CYP450 Inhibition: Understanding Drug-Drug Interactions Due to Mechanism-Based Inhibition in Clinical Practice Malavika Deodhar 1, Sweilem B Al Rihani 1 , Meghan J. Arwood 1, Lucy Darakjian 1, Pamela Dow 1 , Jacques Turgeon 1,2 and Veronique Michaud 1,2,* 1 Tabula Rasa HealthCare Precision Pharmacotherapy Research and Development Institute, Orlando, FL 32827, USA; [email protected] (M.D.); [email protected] (S.B.A.R.); [email protected] (M.J.A.); [email protected] (L.D.); [email protected] (P.D.); [email protected] (J.T.) 2 Faculty of Pharmacy, Université de Montréal, Montreal, QC H3C 3J7, Canada * Correspondence: [email protected]; Tel.: +1-856-938-8697 Received: 5 August 2020; Accepted: 31 August 2020; Published: 4 September 2020 Abstract: In an ageing society, polypharmacy has become a major public health and economic issue. Overuse of medications, especially in patients with chronic diseases, carries major health risks. One common consequence of polypharmacy is the increased emergence of adverse drug events, mainly from drug–drug interactions. The majority of currently available drugs are metabolized by CYP450 enzymes. Interactions due to shared CYP450-mediated metabolic pathways for two or more drugs are frequent, especially through reversible or irreversible CYP450 inhibition. The magnitude of these interactions depends on several factors, including varying affinity and concentration of substrates, time delay between the administration of the drugs, and mechanisms of CYP450 inhibition. Various types of CYP450 inhibition (competitive, non-competitive, mechanism-based) have been observed clinically, and interactions of these types require a distinct clinical management strategy. This review focuses on mechanism-based inhibition, which occurs when a substrate forms a reactive intermediate, creating a stable enzyme–intermediate complex that irreversibly reduces enzyme activity.
    [Show full text]
  • Spring 2013 Lecture 13-14
    CHM333 LECTURE 13 – 14: 2/13 – 15/13 SPRING 2013 Professor Christine Hrycyna INTRODUCTION TO ENZYMES • Enzymes are usually proteins (some RNA) • In general, names end with suffix “ase” • Enzymes are catalysts – increase the rate of a reaction – not consumed by the reaction – act repeatedly to increase the rate of reactions – Enzymes are often very “specific” – promote only 1 particular reaction – Reactants also called “substrates” of enzyme catalyst rate enhancement non-enzymatic (Pd) 102-104 fold enzymatic up to 1020 fold • How much is 1020 fold? catalyst time for reaction yes 1 second no 3 x 1012 years • 3 x 1012 years is 500 times the age of the earth! Carbonic Anhydrase Tissues ! + CO2 +H2O HCO3− +H "Lungs and Kidney 107 rate enhancement Facilitates the transfer of carbon dioxide from tissues to blood and from blood to alveolar air Many enzyme names end in –ase 89 CHM333 LECTURE 13 – 14: 2/13 – 15/13 SPRING 2013 Professor Christine Hrycyna Why Enzymes? • Accelerate and control the rates of vitally important biochemical reactions • Greater reaction specificity • Milder reaction conditions • Capacity for regulation • Enzymes are the agents of metabolic function. • Metabolites have many potential pathways • Enzymes make the desired one most favorable • Enzymes are necessary for life to exist – otherwise reactions would occur too slowly for a metabolizing organis • Enzymes DO NOT change the equilibrium constant of a reaction (accelerates the rates of the forward and reverse reactions equally) • Enzymes DO NOT alter the standard free energy change, (ΔG°) of a reaction 1. ΔG° = amount of energy consumed or liberated in the reaction 2.
    [Show full text]
  • The Crotonase Superfamily: Divergently Related Enzymes That Catalyze Different Reactions Involving Acyl Coenzyme a Thioesters
    Acc. Chem. Res. 2001, 34, 145-157 similar three-dimensional architectures. In each protein, The Crotonase Superfamily: a common structural strategy is employed to lower the Divergently Related Enzymes free energies of chemically similar intermediates. Catalysis of the divergent chemistries is accomplished by both That Catalyze Different retaining those functional groups that catalyze the com- mon partial reaction and incorporating new groups that Reactions Involving Acyl direct the intermediate to new products. Indeed, as a Coenzyme A Thioesters specific example, the enolase superfamily has served as a paradigm for the study of catalytically diverse superfami- HAZEL M. HOLDEN,*,³ lies.3 The active sites of proteins in the enolase superfamily MATTHEW M. BENNING,³ are located at the interfaces between two structural TOOMAS HALLER,² AND JOHN A. GERLT*,² motifs: the catalytic groups are positioned in conserved Departments of Biochemistry, University of Illinois, regions at the ends of the â-strands forming (R/â) 8-barrels, Urbana, Illinois 61801, and University of Wisconsin, while the specificity determinants are found in flexible Madison, Wisconsin 53706 loops in the capping domains formed by the N- and Received August 9, 2000 C-terminal portions of the polypeptide chains. While the members of the enolase superfamily share similar three- ABSTRACT dimensional architectures, they catalyze different overall Synergistic investigations of the reactions catalyzed by several reactions that share a common partial reaction: abstrac- members of an enzyme superfamily provide a more complete tion of an R-proton from a carboxylate anion substrate understanding of the relationships between structure and function than is possible from focused studies of a single enzyme alone.
    [Show full text]
  • ALLOSTERIC REGULATION of PROTHROMBIN ACTIVATION by FACTOR Va
    ALLOSTERIC REGULATION OF PROTHROMBIN ACTIVATION BY FACTOR Va MAHESHEEMA ALI Bachelor of Science in Botany, Microbiology, Chemistry Osmania University, India April 2000 Master of Science in Organic Chemistry Osmania University, India April 2002 Submitted in partial fulfillment of requirements for the degree DOCTOR OF PHILOSOPHY IN CLINICAL AND BIOANALYTICALCHEMISTRY at the CLEVELAND STATE UNIVERITY May 2016 We hereby approve this dissertation for Mahesheema Ali Candidate for the Doctor of Philosophy in Clinical-Bioanalytical Chemistry Degree for the Department of Chemistry and the CLEVELAND STATE UNIVERSITY College of Graduate Studies ________________________________________ Dissertation Chairperson, Dr. Michael Kalafatis Department of CHEMISTRY ______________________ Date ________________________________________ Dissertation Committee Member, Dr. Edward F. Plow Department of MOLECULAR CARDIOLOGY Cleveland Clinic Foundation ______________________ Date ________________________________________ Dr. Anton A. Komar Department of BIOLOGY ______________________ Date ________________________________________ Dr. David J. Anderson Department of CHEMISTRY ______________________ Date ________________________________________ Dr. Crystal M. Weyman Department of BIOLOGY ______________________ Date Date of Defense: April 22nd, 2016 DEDICATION I dedicate this thesis to my loving family. My beloved husband, Mir Ali, has shown unwavering support and encouragement during past five years of my doctoral journey. I am grateful for his love and continuous support, which gave me the strength and courage to pursue my dream and to make it come true. I would like to give special thanks to my wonderful children--Maaz, Maryum, and Idris--for their support and patience throughout. I would like to thank my parents, Mrs. Kaneez Fathima and Mr.Yousuf Ali for all that I have become today, for the constant support in my academic career and personal life.
    [Show full text]
  • An Introduction to Metabolism
    CAMPBELL BIOLOGY IN FOCUS URRY • CAIN • WASSERMAN • MINORSKY • REECE 6 An Introduction to Metabolism Lecture Presentations by Kathleen Fitzpatrick and Nicole Tunbridge, Simon Fraser University © 2016 Pearson Education, Inc. SECOND EDITION The Energy of Life . The living cell is a miniature chemical factory where thousands of reactions occur . The cell extracts energy and applies energy to perform work . Some organisms even convert energy to light, as in bioluminescence © 2016 Pearson Education, Inc. Figure 6.1 © 2016 Pearson Education, Inc. Concept 6.1: An organism’s metabolism transforms matter and energy . Metabolism is the totality of an organism’s chemical reactions . Metabolism is an emergent property of life that arises from interactions between molecules within the cell © 2016 Pearson Education, Inc. Metabolic Pathways . A metabolic pathway begins with a specific molecule and ends with a product . Each step is catalyzed by a specific enzyme © 2016 Pearson Education, Inc. Figure 6.UN01 Enzyme 1 Enzyme 2 Enzyme 3 A B C D Reaction 1 Reaction 2 Reaction 3 Starting Product molecule © 2016 Pearson Education, Inc. Catabolic pathways release energy by breaking down complex molecules into simpler compounds . One example of catabolism is cellular respiration, the breakdown of glucose and other organic fuels to carbon dioxide and water © 2016 Pearson Education, Inc. Anabolic pathways consume energy to build complex molecules from simpler ones . The synthesis of proteins from amino acids is an example of anabolism . Bioenergetics is the study of how energy flows through living organisms © 2016 Pearson Education, Inc. Forms of Energy . Energy is the capacity to cause change . Energy exists in various forms, some of which can perform work © 2016 Pearson Education, Inc.
    [Show full text]
  • Glucokinase Regulatory Protein As the Allosteric Switch for Glucokinase
    Molecular basis for the role of glucokinase regulatory protein as the allosteric switch for glucokinase Jung Min Choia,1, Moon-Hyeong Seoa,1, Hyun-Ho Kyeonga, Eunkyung Kima,2, and Hak-Sung Kima,b,3 aDepartment of Biological Sciences and bGraduate School of Nanoscience and Technology, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea Edited* by Stephen J. Benkovic, Pennsylvania State University, University Park, PA, and approved May 6, 2013 (received for review January 9, 2013) Glucokinase (GK) is a monomeric allosteric enzyme and plays between GK and GKRP and a modulation through the effectors a pivotal role in blood glucose homeostasis. GK is regulated by GK (17, 18). Despite the intensive studies on GK and GKRP, the regulatory protein (GKRP), and indirectly by allosteric effectors of molecular basis for the allosteric regulation mechanism of GK by GKRP. Despite the critical roles of GK and GKRP, the molecular basis GKRP remains poorly understood because of the lack of struc- for the allosteric regulation mechanism of GK by GKRP remains tural information of the GK/GKRP complex. unclear. We determined the crystal structure of Xenopus GK and Here, to demonstrate the molecular mechanism for the allo- GKRP complex in the presence of fructose-6-phosphate at 2.9 Å. steric regulation of GK by GKRP and effectors, we determined GKRP binds to a super-open conformation of GK mainly through the crystal structure of a Xenopus laevis GK and GKRP complex hydrophobic interaction, inhibiting the GK activity by locking a small in the presence of F6P. Structural analysis of the complex and domain of GK.
    [Show full text]
  • Proteolytic Cleavage—Mechanisms, Function
    Review Cite This: Chem. Rev. 2018, 118, 1137−1168 pubs.acs.org/CR Proteolytic CleavageMechanisms, Function, and “Omic” Approaches for a Near-Ubiquitous Posttranslational Modification Theo Klein,†,⊥ Ulrich Eckhard,†,§ Antoine Dufour,†,¶ Nestor Solis,† and Christopher M. Overall*,†,‡ † ‡ Life Sciences Institute, Department of Oral Biological and Medical Sciences, and Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia V6T 1Z4, Canada ABSTRACT: Proteases enzymatically hydrolyze peptide bonds in substrate proteins, resulting in a widespread, irreversible posttranslational modification of the protein’s structure and biological function. Often regarded as a mere degradative mechanism in destruction of proteins or turnover in maintaining physiological homeostasis, recent research in the field of degradomics has led to the recognition of two main yet unexpected concepts. First, that targeted, limited proteolytic cleavage events by a wide repertoire of proteases are pivotal regulators of most, if not all, physiological and pathological processes. Second, an unexpected in vivo abundance of stable cleaved proteins revealed pervasive, functionally relevant protein processing in normal and diseased tissuefrom 40 to 70% of proteins also occur in vivo as distinct stable proteoforms with undocumented N- or C- termini, meaning these proteoforms are stable functional cleavage products, most with unknown functional implications. In this Review, we discuss the structural biology aspects and mechanisms
    [Show full text]