Insights Into the Mechanism of Copper-Tolerance in Fibroporia Radiculosa: the Biosynthesis of Oxalate

Total Page:16

File Type:pdf, Size:1020Kb

Insights Into the Mechanism of Copper-Tolerance in Fibroporia Radiculosa: the Biosynthesis of Oxalate International Biodeterioration & Biodegradation 105 (2015) 90e96 Contents lists available at ScienceDirect International Biodeterioration & Biodegradation journal homepage: www.elsevier.com/locate/ibiod Insights into the mechanism of copper-tolerance in Fibroporia radiculosa: The biosynthesis of oxalate * Katie M. Ohno a, , Carol A. Clausen a, Frederick Green III a, Susan V. Diehl b a USDA Forest Products Laboratory, One Gifford Pinchot Drive, Madison, WI 53726, USA b Mississippi State University, Department of Forest Products, 201 Locksley Way, Starkville, MS 39759, USA article info abstract Article history: Copper is currently used as the key component in wood preservatives despite the known tolerance of Received 18 May 2015 many brown-rot Basidiomycetes. Copper-tolerant fungi, like Fibroporia radiculosa, produce and accu- Received in revised form mulate high levels of oxalate when exposed to copper. To gain insight into the mechanism of oxalate 12 August 2015 production, four F. radiculosa isolates decaying untreated and 1.2% ammoniacal copper citrate-treated Accepted 24 August 2015 wood were evaluated for the differential expression of citrate synthase, isocitrate lyase, glyoxylate de- Available online xxx hydrogenase, a succinate/fumarate antiporter, and a copper resistance-associated ATPase pump. Samples were analyzed at 2, 4, 6, and 8 weeks for oxalate production and gene expression. ATPase pump Keywords: Copper-tolerance expression increased in the presence of copper when initial oxalate concentrations were low, suggesting Brown-rot decay it functions in helping the fungus adapt to the copper-rich environment by pumping toxic copper ions Oxalate out of the cell. A connection in expression levels between citrate synthase, the succinate/fumarate Fibroporia antiporter isocitrate lyase, and glyoxylate dehydrogenase for the four isolates was found suggesting the Antrodia production of oxalate originates in the mitochondrial TCA cycle via citrate synthase, shunts to the Gene expression glyoxysomal glyoxylate cycle via the succinate/fumarate antiporter, moves through a portion of the glyoxylate cycle (isocitrate lyase), and ultimately is made in the cytoplasm (gyloxylate dehydrogenase). Published by Elsevier Ltd. 1. Introduction in decayed wood treated with copper-based preservatives (Freeman and McIntyre, 2008). Another important factor related to The mechanism of copper-tolerance in brown-rot fungi has copper tolerance is the presence of a low pH environment, which is often been associated with the production of oxalate (Murphy and also stimulated by oxalate production (Clausen et al., 2000; Humar Levy, 1983; Sutter et al., 1983; Daniel, 1994; Leithoff et al., 1995; et al., 2001; Clausen and Green, 2003; Green and Clausen, 2003, Pohleven et al., 2002; Green and Clausen, 2003, 2005; Hastrup 2005). Choi et al. (2002) showed spores from copper-tolerant or- et al., 2006; Freeman and McIntyre, 2008; Arango et al., 2009; ganisms did not germinate in the presence of copper because the Schilling and Inda, 2011). High extracellular accumulation of oxa- spores lack the ability to actively produce oxalate (Freeman and late initiates the precipitation of copper into the insoluble form of McIntyre, 2008). Thus, oxalate accumulation and precipitation of copper oxalate forming crystals and renders the copper ion inert copper oxalate crystals play an important role in the mechanism of (Pohleven et al., 1999; De Groot and Woodward, 1999; Humar et al., copper tolerance. 2002). Both copper oxalate and calcium oxalate crystals are present The production of oxalate occurs by numerous classes of fungi and many accumulate it in high concentrations. The majority of brown-rot fungi produce oxalate in detectable amounts; however, Abbreviations: CS, citrate synthase; ICL, isocitrate lyase; GLOXDH, glyoxylate its accumulation is limited in white-rot fungi most likely due to dehydrogenase; ANTI, succinate/fumarate antiporter; ATPase, copper resistance- oxalate decarboxylase (Hastrup et al., 2006). In white-rot fungi, associated ATPase pump; TCA, tricarboxylic acid cycle; GLOX, glyoxylate cycle; intracellular oxalate decarboxylase breaks down oxalate to carbon OXA, oxaloacetase; UN, untreated wood; CC, 1.2% ammoniacal copper citrate treated dioxide and formate (Espejo and Agosin, 1991; Hastrup et al., 2006). wood; RH, relative humidity; SP, Southern pine; CDS, coding sequence; RT-qPCR, In brown-rot fungi, oxalate has the potential to accumulate as a Reverse Transcriptase Quantitative Polymerase Chain Reaction; HK, actin; CT, threshold values. function of fungal respiration (Schilling and Jellison, 2005). Some * Corresponding author. brown-rot fungi produce oxalate for initial decay but do not E-mail address: [email protected] (K.M. Ohno). http://dx.doi.org/10.1016/j.ibiod.2015.08.016 0964-8305/Published by Elsevier Ltd. K.M. Ohno et al. / International Biodeterioration & Biodegradation 105 (2015) 90e96 91 þ accumulate significant levels during the decay process, while other the cytoplasm and functions to pump copper ions (Cu2 ) out of the brown-rot fungi utilize oxalate throughout the decay process cell (Fig. 1). resulting in steady levels of accumulation (Goodell, 2003). Co-biocides are needed to prevent these copper-tolerant or- Several pathways have been proposed for the biosynthesis of ganisms from circumventing the copper-based preventative mea- oxalate in brown-rot fungi; however, in recent years, many re- sures in use today; therefore, it is critical that these mechanisms be searchers have linked it to the tricarboxylic acid (TCA) and glyox- investigated. Knowing the exact mechanisms and specific re- ylate (GLOX) cycles (Dutton and Evans, 1996; Munir et al., 2000; quirements of organisms involved in copper tolerance would give Munir et al., 2001; Yoon et al., 2002; Schilling and Jellison, 2005; insight into the physiological intricacies of copper tolerance. A Hastrup et al., 2006). The TCA cycle is localized in the mitochon- better understanding of this mechanism is crucial in developing dria while the GLOX cycle takes place in the glyoxysome (Dutton targeted methods to control these particular organisms. The pur- and Evans, 1996). Munir et al., (2001) investigated oxalate biosyn- pose of this study is to gain insight into the biosynthesis of oxalate thesis in the copper-tolerant fungus Tyromyces palustris. This study by which the brown-rot decay fungus, F. radiculosa, regulates hypothesized that 1 mol of glucose is converted to 2 mol of oxalic tolerance of copper-treated wood. Specific objectives of this study acid rather than producing the traditional CO2 from the TCA cycle. are to examine select F. radiculosa isolates during decay of un- Through their work, it was discovered that there are two major treated and copper-treated wood for the differential expression of oxalate producing enzymes, oxaloacetase (OXA) and glyoxylate genes associated with oxalate production, and to propose a po- dehydrogenase (GLOXDH). Munir et al. (2001) discovered quanti- tential pathway involved in the production of oxalate. fiable OXA and GLOXDH enzymatic activity, with OXA synthesizing more oxalate than GLOXDH in T. palustris. 2. Materials and methods Using this discovery as a model, we hypothesized that Fibroporia radiculosa, another copper-tolerant fungus, may use a similar 2.1. Sample preparation mechanism for oxalate production. Initial enzymatic analysis showed quantifiable GLOXDH, but not OXA, in four isolates of F. Four isolates of F. radiculosa (Peck) Gilb. & Ryvarden were used radiculosa (unpublished data). These results were related to the in this study: FP-90848-T, L-9414-SP, L-11659-SP, and TFFH 294 genome analysis of F. radiculosa isolate TFFH 294 showing up- (USDA Forest Products Laboratory, Madison, WI). Cultures were regulation of GLOXDH, not OXA, in the early stages of decay maintained on 2% Malt Extract Agar (BD, Fisher Scientific). Isolates (Tang, 2011). In addition to GLOXDH, Tang et al. (2012) found up- were subjected to an AWPA Standard E10-15 (2015) decay chamber regulation of citrate synthase (CS), a succinate-fumarate anti- at 26.7 C and 70% relative humidity (RH) for 2 weeks to allow porter (ANTI), isocitrate lyase (ICL), and a copper resistance- initial colonization (AWPA, 2015). Southern pine (SP) test wafers associated ATPase pump (ATPase) during the early stages of measuring 70 mm  22 mm  4mm(t r  l) were vacuum decay. Citrate synthase is found in the mitochondria as part of the treated with 1.2% ammoniacal copper citrate (CC) for 40 min TCA cycle and converts oxaloacetate to citrate (Fig. 1). The succi- at À172 kPa. After initial colonization, two CC-treated SP test wafers nate/fumarate antiporter, found in the cytoplasm, functions as a were placed directly on top of the colonized feeder strips and shuttle from isocitrate in the TCA cycle to isocitrate in the GLOX incubated at 26.7 C 70%RH for 8 weeks. Untreated SP test wafers cycle which is located in the glyoxysome. The ATPase pump is not were also exposed to the four isolates and incubated at 26.7 C, 70% associated with either the TCA or GLOX cycles; instead, it is found in RH for 8 weeks. Both untreated and CC-treated SP test wafers Fig. 1. Genes (italics) up-regulated in early decay. (A) TCA cycle; (B) GLOX cycle. Light gray box indicates TCA cycle in the mitochondria; dark gray box indicates GLOX cycle in the glyoxysome (Tang, 2011). 92 K.M. Ohno et al. / International Biodeterioration & Biodegradation 105 (2015) 90e96 exposed to the four isolates were harvested at weeks
Recommended publications
  • Proteomic Analysis of Mycelial Proteins from Magnaporthe Oryzae Under Nitrogen Starvation
    Proteomic analysis of mycelial proteins from Magnaporthe oryzae under nitrogen starvation X.-G. Zhou1,2, P. Yu 2, C. Dong2, C.-X. Yao2, Y.-M. Ding2, N. Tao2 and Z.-W. Zhao1 1State Key Laboratory for Conservation and Utilization of Bio-Resources in Yunnan, Yunnan University, Kunming, China 2Key Laboratory of Southwestern Crop Gene Resources and Germplasm Innovation, Ministry of Agriculture and Yunnan Provincial Key Laboratory of Agricultural Biotechnology, Biotechnology and Germplasm Resources Institute, Yunnan Academy of Agricultural Sciences, Kunming, China Corresponding author: Z.W. Zhao E-mail: [email protected] Genet. Mol. Res. 15 (2): gmr.15028637 Received March 23, 2016 Accepted April 11, 2016 Published May 13, 2016 DOI http://dx.doi.org/10.4238/gmr.15028637 ABSTRACT. Magnaporthe oryzae is an important model system in studies of plant pathogenic fungi, and nitrogen is a key nutrient source affecting microbial growth and development. In order to understand how nitrogen stress causes changes in mycelial proteins, we analyzed differentially expressed mycelial proteins from the M. oryzae virulent strain CH-63 using two-dimensional electrophoresis and mass spectrometry in complete medium or under nitrogen starvation conditions. A total of 975 ± 70 and 1169 ± 90 protein spots were detected in complete medium and under nitrogen starvation conditions, respectively. Forty-nine protein spots exhibited at least 2-fold up- regulation or down-regulation at the protein level according to PDQuest7.4. Moreover, 43 protein spots were successfully identified by matrix-assisted laser desorption/ionization-time-of-flight/time-of-flight mass spectrometry. Among these spots, 6 proteins were functionally unknown and 37 proteins were categorized into 5 groups according to Genetics and Molecular Research 15 (2): gmr.15028637 ©FUNPEC-RP www.funpecrp.com.br X.-G.
    [Show full text]
  • Oxdc Antibody Rabbit Polyclonal Antibody Catalog # ABV11223
    10320 Camino Santa Fe, Suite G San Diego, CA 92121 Tel: 858.875.1900 Fax: 858.622.0609 OxdC Antibody Rabbit Polyclonal Antibody Catalog # ABV11223 Specification OxdC Antibody - Product Information Application WB Primary Accession O34714 Reactivity Human Host Rabbit Clonality Polyclonal Isotype Rabbit IgG Calculated MW 43566 OxdC Antibody - Additional Information Gene ID 938620 Positive Control Western Blot: Recombinant protein Application & Usage Western blot: 1-4 Western blot of Oxalate decarboxylase µg/ml. antibody. Lane 1: rb- Oxalate decarboxylase - Other Names 10 ng. Lane 2: rb- Oxalate decarboxylase - 50 YvrK ng Target/Specificity OxdC OxdC Antibody - Background Antibody Form Oxalate decarboxylase (OxdC, EC4.1.1.2) is a Liquid manganese-containing enzyme, which decomposes oxalic acid and oxalate. With Appearance OxdC catalysis, oxalate is split into formate Colorless liquid and CO2. This enzyme belongs to the family of lyases, specifically the carboxy-lyases, which Formulation 100 µg (0.5 mg/ml) of antibody in PBS pH cleave carbon-carbon bonds. The systematic 7.2 containing 0.01 % BSA, 0.01 % name of this enzyme class is oxalate thimerosal, and 50 % glycerol. carboxy-lyase (formate-forming). This enzyme is also called oxalate carboxy-lyase. The Handling enzyme is composed of two cupin domains, The antibody solution should be gently each of which contains a Mn (II) ion. This mixed before use. enzyme participates in glyoxylate and dicarboxylate metabolism. This enzyme has Reconstitution & Storage been recognized for diagnostics in diverse -20 °C biotechnological applications such as the clinical assay of oxalate in blood and urine, Background Descriptions therapeutics, process industry, and agriculture to lower oxalate levels in foods and the environment.
    [Show full text]
  • Proteo-Metabolomic Investigation of Transgenic Rice Unravels Metabolic
    www.nature.com/scientificreports OPEN Proteo-metabolomic investigation of transgenic rice unravels metabolic alterations and Received: 27 November 2018 Accepted: 24 June 2019 accumulation of novel proteins Published: xx xx xxxx potentially involved in defence against Rhizoctonia solani Subhasis Karmakar1, Karabi Datta1, Kutubuddin Ali Molla2,3, Dipak Gayen4, Kaushik Das1, Sailendra Nath Sarkar1 & Swapan K. Datta1 The generation of sheath blight (ShB)-resistant transgenic rice plants through the expression of Arabidopsis NPR1 gene is a signifcant development for research in the feld of biotic stress. However, to our knowledge, regulation of the proteomic and metabolic networks in the ShB-resistant transgenic rice plants has not been studied. In the present investigation, the relative proteome and metabolome profles of the non–transformed wild-type and the AtNPR1-transgenic rice lines prior to and subsequent to the R. solani infection were investigated. Total proteins from wild type and transgenic plants were investigated using two-dimensional gel electrophoresis (2-DE) followed by mass spectrometry (MS). The metabolomics study indicated an increased abundance of various metabolites, which draws parallels with the proteomic analysis. Furthermore, the proteome data was cross-examined using network analysis which identifed modules that were rich in known as well as novel immunity-related prognostic proteins, particularly the mitogen-activated protein kinase 6, probable protein phosphatase 2C1, probable trehalose-phosphate phosphatase 2 and heat shock protein. A novel protein, 14–3– 3GF14f was observed to be upregulated in the leaves of the transgenic rice plants after ShB infection, and the possible mechanistic role of this protein in ShB resistance may be investigated further.
    [Show full text]
  • Oxalate Decarboxylase from Bacillus Subtilis, Recombinant Cat
    Oxalate Decarboxylase from Bacillus subtilis, recombinant Cat. No. NATE-1688 Lot. No. (See product label) Introduction Description Oxalate decarboxylase (OxdC, EC4.1.1.2) is a manganese-containing enzyme, which decomposes oxalic acid and oxalate. With OxdC catalysis, oxalate is split into formate and CO2. This enzyme belongs to the family of lyases, specifically the carboxy-lyases, which cleave carbon-carbon bonds. The systematic name of this enzyme class is oxalate carboxy-lyase (formate-forming). This enzyme is also called oxalate carboxy-lyase. The enzyme is composed of two cupin domains, each of which contains a Mn (II) ion. This enzyme participates in glyoxylate and dicarboxylate metabolism. This enzyme has been recognized for diagnostics in diverse biotechnological applications such as the clinical assay of oxalate in blood and urine, therapeutics, process industry, and agriculture to lower oxalate levels in foods and the environment. The recombinant protein made from the Bacillus Subtilis sequence includes OxdC with N-terminal His-tag. Synonyms Oxalate carboxy-lyase; EC 4.1.1.2; Oxalate decarboxylase; OxdC Product Information Species Bacillus subtilis Source E. coli Form Liquid Formulation In 50 mM NaOAC, pH 5.5. The activation was stopped by addition of 10 mM MMTS, which can be removed under reducing conditions. EC Number EC 4.1.1.2 CAS No. 9024-97-9 Molecular Weight 45.9 kDa Purity > 98% by SDS-PAGE Activity 150U/mg Creative Enzymes. All rights reserved. 45-1 Ramsey Road, Shirley, NY 11967, USA Tel:1-631-562-8517 1-516-512-3133 Fax:1-631-938-8127 E-mail: [email protected] http://www.creative-enzymes.com Concentration 2 mg/mL Unit Definition One unit is the amount of enzyme that generates 1.0 µmole of NADH at 37°C.
    [Show full text]
  • The Structure of Oxalate Decarboxylase at Its Active Ph
    bioRxivRunning preprint head: doi: https://doi.org/10.1101/426874 Low pH Structure, Oxalate; this version Decarboxylase posted September 25, 2018. The copyright holder for this preprint (which was1 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The Structure of Oxalate Decarboxylase at its Active pH M. J. Burg, J. L. Goodsell, U. T. Twahir, S. D. Bruner, and A. Angerhofer, bioRxivLow preprintpH Structure, doi: https://doi.org/10.1101/426874 Oxalate Decarboxylase; this version posted September 25, 2018. The copyright holder for this preprint (which was2 not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Abstract Oxalate decarboxylase catalyzes the redox-neutral unimolecular disproportionation reaction of oxalic acid. The pH maximum for catalysis is ~4.0 and activity is negligible above pH7. Here we report on the first crystal structure of the enzyme in its active pH range at pH4.6, and at a resolution of 1.45 Å, the highest to date. The fundamental tertiary and quaternary structure of the enzyme does not change with pH. However, the low pH crystals are heterogeneous containing both a closed and open conformation of a flexible loop region which gates access to the N-terminal active site cavity. Residue E162 in the closed conformation points away from the active-site Mn ion owing to the coordination of a buffer molecule, acetate. Since the quaternary structure of the enzyme appears unaffected by pH many conclusions drawn from the structures taken at high pH remain valid.
    [Show full text]
  • MECHANISM of the REACTION CATALYZED by the OXALATE DECARBOXYLASE from Bacillus Subtilis
    MECHANISM OF THE REACTION CATALYZED BY THE OXALATE DECARBOXYLASE FROM Bacillus subtilis By DRAŽENKA SVEDRUŽIĆ A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2005 Copyright 2005 by Draženka Svedružić This thesis is dedicated to my brother Željko, my parents and Chris. Ova je teza posvećena mome bratu Željku, mojim roditeljima i Krisu. ACKNOWLEDGMENTS This study was supported by grants from the National Institutes of Health (DK61666 and DK53556) and by the University of Florida Chemistry Department. Partial funding was also received from Dr. Ammon B. Peck. Thanks go to my doctoral dissertation committee: Dr. Steven A. Benner, Dr. Ammon B. Peck, Dr. Michael J. Scott, Dr. Jon D. Stewart, and especially my advisor, Dr. Nigel G. J. Richards, for making this research project a reality, but also for his constant support and guidance. Dr. Laurie A. Renhardt, Yang Liu and Dr. Wallace W. Cleland I thank for fruitful collaboration on heavy atom isotope effects research. Also, I thank Dr. Wallace W. Cleland for hospitality during my stay in Madison, Wisconsin. Thanks go to my EPR collaborators Dr. Lee Walker, Dr. Andrzej Ozarowski and Dr. Alexander Angerhofer. I am grateful to my coworkers and friends in the Richards research group, especially Dr. Christopher H. Chang for proofreading, countless discussions, valuable insights, guidance and support. Special thanks go to Stefan Jonsson for all his help, Sue Abbatiello for her mass spectrometry efforts and Lukas Koroniak for help with NMR experiments. Also thanks go to all members of Richards group, especially Mihai, Jemy and Cory, for providing a pleasant and supporting environment in and out of lab.
    [Show full text]
  • (12) Patent Application Publication (10) Pub. No.: US 2009/0292100 A1 Fiene Et Al
    US 20090292100A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2009/0292100 A1 Fiene et al. (43) Pub. Date: Nov. 26, 2009 (54) PROCESS FOR PREPARING (86). PCT No.: PCT/EP07/57646 PENTAMETHYLENE 1.5-DIISOCYANATE S371 (c)(1), (75) Inventors: Martin Fiene, Niederkirchen (DE): (2), (4) Date: Jan. 9, 2009 (DE);Eckhard Wolfgang Stroefer, Siegel, Mannheim (30) Foreign ApplicationO O Priority Data Limburgerhof (DE); Stephan Aug. 1, 2006 (EP) .................................. O61182.56.4 Freyer, Neustadt (DE); Oskar Zelder, Speyer (DE); Gerhard Publication Classification Schulz, Bad Duerkheim (DE) (51) Int. Cl. Correspondence Address: CSG 18/00 (2006.01) OBLON, SPIVAK, MCCLELLAND MAIER & CD7C 263/2 (2006.01) NEUSTADT, L.L.P. CI2P I3/00 (2006.01) 194O DUKE STREET CD7C 263/10 (2006.01) ALEXANDRIA, VA 22314 (US) (52) U.S. Cl. ........... 528/85; 560/348; 435/128; 560/347; 560/355 (73) Assignee: BASFSE, LUDWIGSHAFEN (DE) (57) ABSTRACT (21) Appl. No.: 12/373,088 The present invention relates to a process for preparing pen tamethylene 1,5-diisocyanate, to pentamethylene 1,5-diiso (22) PCT Filed: Jul. 25, 2007 cyanate prepared in this way and to the use thereof. US 2009/0292100 A1 Nov. 26, 2009 PROCESS FOR PREPARING ene diisocyanates, especially pentamethylene 1,4-diisocyan PENTAMETHYLENE 1.5-DIISOCYANATE ate. Depending on its preparation, this proportion may be up to several % by weight. 0014. The pentamethylene 1,5-diisocyanate prepared in 0001. The present invention relates to a process for pre accordance with the invention has, in contrast, a proportion of paring pentamethylene 1,5-diisocyanate, to pentamethylene the branched pentamethylene diisocyanate isomers of in each 1.5-diisocyanate prepared in this way and to the use thereof.
    [Show full text]
  • Comparative Studies of Oxalyl-Coa Decarboxylase Produced by Soil
    3t' (O' COMPARATIVE STUDIES OF OXALYL.COA DECARBOXYLASE PRODUCED BY SOIL AND RUMINAL BACTERIA Thesis Submitted for the degree of Master of Agricultural Science in The University of Adelaide Faculty of Agricultural and Natural Resource Sciences by STEPHEN BOTTRILL November 1999 I I Table of Contents List of Figures VI List of Tables VM Abstract IX Acknowledgements XII Ståtement XIII List of Abbreviations XTV Chapær 1. Liærature Review 1 1.1 Introduction. 1 1.2 Exogenous Sources of Oxalates. 1 1.3 Endogenous Sources of Oxalate. 5 1.4 Poisoning. 9 1.4.1 Acute Poisoning" 10 1.4.2 Subacute Poisoning. 11 1.4.3 Chronic Poisoning. t2 1.4.4 SymPtoms in Humans. 14 1.4.5 Treatment of Poisoning. I4 1.4.6 Management to Prevent Poisoning. 15 1.5 Oxalate-Degrading Microorganisms. 18 1.6 Bacterial Classification 22 1.7 Pathways of Oxalate Degradation. 24 1.8 Formate in the Rumen. 27 1.9 Aims and Objectives. 29 Chapær 2. Materials and Methods 31 2.1 Materials 31 2.1.1 Chemicals 3r 2.1.2 EquiPment 31 2.I.3 Bacterial Strains and Plasmids 32 TI 2.1.4 Composition of Media 34 2.1.4.I Oxalate-Containing Media 34 2.1.4.I.1Liquid 34 2.1.4.1.2 Solid 34 2.1.4.2 O mlob act er formi g enes Media 35 2.I.4.2.I Trace Metals Solution 35 2.I.4.2.2 Medium A 35 2.1.4.2.3 Medium B 36 2.I.4.3 Luria-Bertani (LB) Broth 36 2.I.4.4 SOC Medium 37 2.2 Methods 37 2.2 -I Growth conditions 37 2.2.2 Isolation of oxal ate- de gradin g s oil bacteria 37 2.2.3 Characterisation of soil isolaæs 38 2.2.3.1 MicroscoPY 38 2.2.3.2 Gram stain 38 2.2.3.3 Carbon source utilisation 40 2.2.4.4 Volatile
    [Show full text]
  • Structural and Mechanistic Studies on Α-Amino Β-Carboxymuconate Ε
    Georgia State University ScholarWorks @ Georgia State University Chemistry Dissertations Department of Chemistry Summer 8-12-2014 Structural and Mechanistic Studies on α-Amino β- Carboxymuconate ε-Semialdehyde Decarboxylase and α- Aminomuconate ε-Semialdehyde Dehydrogenase Lu Huo Georgia State University Follow this and additional works at: https://scholarworks.gsu.edu/chemistry_diss Recommended Citation Huo, Lu, "Structural and Mechanistic Studies on α-Amino β-Carboxymuconate ε-Semialdehyde Decarboxylase and α-Aminomuconate ε-Semialdehyde Dehydrogenase." Dissertation, Georgia State University, 2014. https://scholarworks.gsu.edu/chemistry_diss/100 This Dissertation is brought to you for free and open access by the Department of Chemistry at ScholarWorks @ Georgia State University. It has been accepted for inclusion in Chemistry Dissertations by an authorized administrator of ScholarWorks @ Georgia State University. For more information, please contact [email protected]. STRUCTURAL AND MECHANISTIC STUDIES ON α-AMINO β-CARBOXYMUCONATE ε-SEMIALDEHYDE DECARBOXYLASE AND α-AMINOMUCONATE ε-SEMIALDEHYDE DEHYDROGENASE by LU HUO Under the Direction of Dr. Aimin Liu ABSTRACT α-Amino-β-carboxymuconate-ε-semialdehyde decarboxylase (ACMSD) and α- aminomuconate-ε-semialdehyde dehydrogenase (AMSDH) are two neighboring enzymes in the L-tryptophan and 2-nitrobenzoic acid degradation pathways. The substrates of the two enzymes, α-amino-β-carboxymuconate-ε-semialdehyde (ACMS) and α-aminomuconate-ε-semialdehyde (2-AMS), are unstable and spontaneously decay to quinolinic acid and picolinic acid, respectively. ACMSD utilizes a divalent zinc metal as cofactor and is a member of the amidohydrolase superfamily. In this dissertation work, we have identified an important histidine residue in the active site that plays dual roles in tuning metal selectivity and activating a metal bound water ligand using mutagenesis, resonance Raman, EPR, crystallography, and ICP metal analysis techniques.
    [Show full text]
  • Crystal Structure of Oxalate Decarboxylase from Photorhabdus Luminescens, a Symbiotic Bacterium Associated with Entomopathogenic Nematodes
    RESEARCH ARTICLES Crystal structure of oxalate decarboxylase from Photorhabdus luminescens, a symbiotic bacterium associated with entomopathogenic nematodes Sreeja Chellappan1,2, S. Mathivanan1, R. Thippeswamy3, M. Nagesh3, H. S. Savithri4 and M. R. N. Murthy1,5,6,* 1Molecular Biophysics Unit and 4Department of Biochemistry, Indian Institute of Science, Bengaluru 560 012, India 2Department of Molecular Biology, Kannur University, Kannur 671 314, India 3Indian Council of Agricultural Research, Project Directorate of Biological Control, Bengaluru 560 024, India 5Institute of Bioinformatics and Applied Biotechnology, Bengaluru 560 100, India 6Indian Institute of Science Education and Research, Thiruvananthapuram 695 551, India promotes nematode development and multiplication. Photorhabdus luminescens is a Gram-negative, symbi- 3 otic bacterium associated with entomopathogenic Duchaud et al. have determined the complete sequence nematodes of the genus Heterorhabditis. Several genes of P. luminescens. Analysis of the genomic sequence from this organism related to insecticidal properties revealed that it possesses a number of genes encoding for are being examined for their potential in pest man- potential virulence factors. agement. Oxalate decarboxylase (OXDC), an enzyme Oxalate decarboxylase (OXDC), proposed to have secreted by bacteria and fungi and putatively asso- insecticidal effect and secreted by bacteria, catalyses the ciated with insecticidal pathways catalyses the man- manganese-dependent decarboxylation of oxalate to ganese dependent decarboxylation of oxalate to formate and CO2. OXDC was first reported from fungi formate and CO2. In this study, we report the X-ray Flammulina (Collybia) velutipes and Coriolus hersutus4. crystal structure of OXDC isolated and purified from Later it was found in several other organisms5–12. OXDC Photorhabdus luminescens (PlOXDC, MW 43 kDa) structures have been determined from Bacillus subtilis determined at 1.97 Å resolution.
    [Show full text]
  • Photorespiration Pathways in a Chemolithoautotroph
    bioRxiv preprint doi: https://doi.org/10.1101/2020.05.08.083683; this version posted May 9, 2020. 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 4.0 International license. Photorespiration pathways in a chemolithoautotroph Nico J. Claassens*1, Giovanni Scarinci*1, Axel Fischer1, Avi I. Flamholz2, William Newell1, Stefan Frielingsdorf3, Oliver Lenz3, Arren Bar-Even†1 1Max Planck Institute of Molecular Plant Physiology, Am Mühlenberg 1, 14476 Potsdam-Golm, Germany 2Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, United States. 3Institut für Chemie, Physikalische Chemie, Technische Universität Berlin, Strasse des 17. Juni 135, 10623 Berlin, Germany †corresponding author; phone: +49 331 567-8910; Email: [email protected] *contributed equally Key words: CO2 fixation; hydrogen-oxidizing bacteria; glyoxylate shunt; malate synthase; oxalate metabolism 1 bioRxiv preprint doi: https://doi.org/10.1101/2020.05.08.083683; this version posted May 9, 2020. 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 4.0 International license. Abstract Carbon fixation via the Calvin cycle is constrained by the side activity of Rubisco with dioxygen, generating 2-phosphoglycolate. The metabolic recycling of 2-phosphoglycolate, an essential process termed photorespiration, was extensively studied in photoautotrophic organisms, including plants, algae, and cyanobacteria, but remains uncharacterized in chemolithoautotrophic bacteria.
    [Show full text]
  • Generate Metabolic Map Poster
    Authors: Zheng Zhao, Delft University of Technology Marcel A. van den Broek, Delft University of Technology S. Aljoscha Wahl, Delft University of Technology Wilbert H. Heijne, DSM Biotechnology Center Roel A. Bovenberg, DSM Biotechnology Center Joseph J. Heijnen, Delft University of Technology An online version of this diagram is available at BioCyc.org. Biosynthetic pathways are positioned in the left of the cytoplasm, degradative pathways on the right, and reactions not assigned to any pathway are in the far right of the cytoplasm. Transporters and membrane proteins are shown on the membrane. Marco A. van den Berg, DSM Biotechnology Center Peter J.T. Verheijen, Delft University of Technology Periplasmic (where appropriate) and extracellular reactions and proteins may also be shown. Pathways are colored according to their cellular function. PchrCyc: Penicillium rubens Wisconsin 54-1255 Cellular Overview Connections between pathways are omitted for legibility. Liang Wu, DSM Biotechnology Center Walter M. van Gulik, Delft University of Technology L-quinate phosphate a sugar a sugar a sugar a sugar multidrug multidrug a dicarboxylate phosphate a proteinogenic 2+ 2+ + met met nicotinate Mg Mg a cation a cation K + L-fucose L-fucose L-quinate L-quinate L-quinate ammonium UDP ammonium ammonium H O pro met amino acid a sugar a sugar a sugar a sugar a sugar a sugar a sugar a sugar a sugar a sugar a sugar K oxaloacetate L-carnitine L-carnitine L-carnitine 2 phosphate quinic acid brain-specific hypothetical hypothetical hypothetical hypothetical
    [Show full text]