Table S2 B Up-Regulated Proteins (Fold Changes) Revealed by Quantitative I TRAQ Proteomic An
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Part One Amino Acids As Building Blocks
Part One Amino Acids as Building Blocks Amino Acids, Peptides and Proteins in Organic Chemistry. Vol.3 – Building Blocks, Catalysis and Coupling Chemistry. Edited by Andrew B. Hughes Copyright Ó 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim ISBN: 978-3-527-32102-5 j3 1 Amino Acid Biosynthesis Emily J. Parker and Andrew J. Pratt 1.1 Introduction The ribosomal synthesis of proteins utilizes a family of 20 a-amino acids that are universally coded by the translation machinery; in addition, two further a-amino acids, selenocysteine and pyrrolysine, are now believed to be incorporated into proteins via ribosomal synthesis in some organisms. More than 300 other amino acid residues have been identified in proteins, but most are of restricted distribution and produced via post-translational modification of the ubiquitous protein amino acids [1]. The ribosomally encoded a-amino acids described here ultimately derive from a-keto acids by a process corresponding to reductive amination. The most important biosynthetic distinction relates to whether appropriate carbon skeletons are pre-existing in basic metabolism or whether they have to be synthesized de novo and this division underpins the structure of this chapter. There are a small number of a-keto acids ubiquitously found in core metabolism, notably pyruvate (and a related 3-phosphoglycerate derivative from glycolysis), together with two components of the tricarboxylic acid cycle (TCA), oxaloacetate and a-ketoglutarate (a-KG). These building blocks ultimately provide the carbon skeletons for unbranched a-amino acids of three, four, and five carbons, respectively. a-Amino acids with shorter (glycine) or longer (lysine and pyrrolysine) straight chains are made by alternative pathways depending on the available raw materials. -
Generated by SRI International Pathway Tools Version 25.0, Authors S
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. Periplasmic (where appropriate) and extracellular reactions and proteins may also be shown. Pathways are colored according to their cellular function. Gcf_000238675-HmpCyc: Bacillus smithii 7_3_47FAA Cellular Overview Connections between pathways are omitted for legibility. -
Characterization of the Human O-Phosphoethanolamine Phospholyase, an Unconventional Pyridoxal Phosphate- Dependent -Lyase
Department of Pharmacy Laboratories of Biochemistry and Molecular Biology PhD Program in Biochemistry and Molecular Biology XXVII cycle Characterization of the human O-phosphoethanolamine phospholyase, an unconventional pyridoxal phosphate- dependent -lyase Coordinator: Prof. Andrea Mozzarelli Tutor: Prof. Alessio Peracchi PhD student: DAVIDE SCHIROLI 2012-2014 1 INDEX: Chapter 1: A subfamily of PLP-dependent enzymes specialized in handling terminal amines Abstract……………………………………………...pg.10 Introduction………………………………………….pg.11 -Nomenclature issues: subgroup-II aminotransferases, class-III ami- notransferases or -aminotransferases?..........................................pg.13 Chemical peculiarities of the reactions catalyzed by AT-II enzymes………………………………………pg.18 - Equilibria in -amine transaminase reactions……………………...pg.18 - Specificity and dual-specificity issues…………………………….…pg.22 Structural peculiarities of AT-II enzymes………...pg.23 - AT-II vs. AT-I enzymes. Comparing the overall structures………..pg.23 - AT-II vs. AT-I. Comparing the PLP-binding sites…………………...pg.25 - The substrate binding site: a gateway system in -KG-specific AT-II transaminases……………………………………………………………pg.31 - The substrate binding site: P and O pockets in pyruvate-specific AT-II transaminases………………………………………………………...….pg.35 - An overview of substrate specificity in AT-II transaminases………pg.42 2 AT-II enzymes that are not aminotransferases……….. …………………………………………………….....pg.43 Inferences on the evolution of AT-II enzymes………… ………………………………………………………..pg.47 Conclusions……………..…………………………..pg.53 -
Supplementary Materials
Supplementary Materials Figure S1. Differentially abundant spots between the mid-log phase cells grown on xylan or xylose. Red and blue circles denote spots with increased and decreased abundance respectively in the xylan growth condition. The identities of the circled spots are summarized in Table 3. Figure S2. Differentially abundant spots between the stationary phase cells grown on xylan or xylose. Red and blue circles denote spots with increased and decreased abundance respectively in the xylan growth condition. The identities of the circled spots are summarized in Table 4. S2 Table S1. Summary of the non-polysaccharide degrading proteins identified in the B. proteoclasticus cytosol by 2DE/MALDI-TOF. Protein Locus Location Score pI kDa Pep. Cov. Amino Acid Biosynthesis Acetylornithine aminotransferase, ArgD Bpr_I1809 C 1.7 × 10−4 5.1 43.9 11 34% Aspartate/tyrosine/aromatic aminotransferase Bpr_I2631 C 3.0 × 10−14 4.7 43.8 15 46% Aspartate-semialdehyde dehydrogenase, Asd Bpr_I1664 C 7.6 × 10−18 5.5 40.1 17 50% Branched-chain amino acid aminotransferase, IlvE Bpr_I1650 C 2.4 × 10−12 5.2 39.2 13 32% Cysteine synthase, CysK Bpr_I1089 C 1.9 × 10−13 5.0 32.3 18 72% Diaminopimelate dehydrogenase Bpr_I0298 C 9.6 × 10−16 5.6 35.8 16 49% Dihydrodipicolinate reductase, DapB Bpr_I2453 C 2.7 × 10−6 4.9 27.0 9 46% Glu/Leu/Phe/Val dehydrogenase Bpr_I2129 C 1.2 × 10−30 5.4 48.6 31 64% Imidazole glycerol phosphate synthase Bpr_I1240 C 8.0 × 10−3 4.7 22.5 8 44% glutamine amidotransferase subunit Ketol-acid reductoisomerase, IlvC Bpr_I1657 C 3.8 × 10−16 -
B Number Gene Name Mrna Intensity Mrna
sample) total list predicted B number Gene name assignment mRNA present mRNA intensity Gene description Protein detected - Membrane protein membrane sample detected (total list) Proteins detected - Functional category # of tryptic peptides # of tryptic peptides # of tryptic peptides detected (membrane b0002 thrA 13624 P 39 P 18 P(m) 2 aspartokinase I, homoserine dehydrogenase I Metabolism of small molecules b0003 thrB 6781 P 9 P 3 0 homoserine kinase Metabolism of small molecules b0004 thrC 15039 P 18 P 10 0 threonine synthase Metabolism of small molecules b0008 talB 20561 P 20 P 13 0 transaldolase B Metabolism of small molecules chaperone Hsp70; DNA biosynthesis; autoregulated heat shock b0014 dnaK 13283 P 32 P 23 0 proteins Cell processes b0015 dnaJ 4492 P 13 P 4 P(m) 1 chaperone with DnaK; heat shock protein Cell processes b0029 lytB 1331 P 16 P 2 0 control of stringent response; involved in penicillin tolerance Global functions b0032 carA 9312 P 14 P 8 0 carbamoyl-phosphate synthetase, glutamine (small) subunit Metabolism of small molecules b0033 carB 7656 P 48 P 17 0 carbamoyl-phosphate synthase large subunit Metabolism of small molecules b0048 folA 1588 P 7 P 1 0 dihydrofolate reductase type I; trimethoprim resistance Metabolism of small molecules peptidyl-prolyl cis-trans isomerase (PPIase), involved in maturation of b0053 surA 3825 P 19 P 4 P(m) 1 GenProt outer membrane proteins (1st module) Cell processes b0054 imp 2737 P 42 P 5 P(m) 5 GenProt organic solvent tolerance Cell processes b0071 leuD 4770 P 10 P 9 0 isopropylmalate -
Supplementary Information
Supplementary information (a) (b) Figure S1. Resistant (a) and sensitive (b) gene scores plotted against subsystems involved in cell regulation. The small circles represent the individual hits and the large circles represent the mean of each subsystem. Each individual score signifies the mean of 12 trials – three biological and four technical. The p-value was calculated as a two-tailed t-test and significance was determined using the Benjamini-Hochberg procedure; false discovery rate was selected to be 0.1. Plots constructed using Pathway Tools, Omics Dashboard. Figure S2. Connectivity map displaying the predicted functional associations between the silver-resistant gene hits; disconnected gene hits not shown. The thicknesses of the lines indicate the degree of confidence prediction for the given interaction, based on fusion, co-occurrence, experimental and co-expression data. Figure produced using STRING (version 10.5) and a medium confidence score (approximate probability) of 0.4. Figure S3. Connectivity map displaying the predicted functional associations between the silver-sensitive gene hits; disconnected gene hits not shown. The thicknesses of the lines indicate the degree of confidence prediction for the given interaction, based on fusion, co-occurrence, experimental and co-expression data. Figure produced using STRING (version 10.5) and a medium confidence score (approximate probability) of 0.4. Figure S4. Metabolic overview of the pathways in Escherichia coli. The pathways involved in silver-resistance are coloured according to respective normalized score. Each individual score represents the mean of 12 trials – three biological and four technical. Amino acid – upward pointing triangle, carbohydrate – square, proteins – diamond, purines – vertical ellipse, cofactor – downward pointing triangle, tRNA – tee, and other – circle. -
Supplementary Informations SI2. Supplementary Table 1
Supplementary Informations SI2. Supplementary Table 1. M9, soil, and rhizosphere media composition. LB in Compound Name Exchange Reaction LB in soil LBin M9 rhizosphere H2O EX_cpd00001_e0 -15 -15 -10 O2 EX_cpd00007_e0 -15 -15 -10 Phosphate EX_cpd00009_e0 -15 -15 -10 CO2 EX_cpd00011_e0 -15 -15 0 Ammonia EX_cpd00013_e0 -7.5 -7.5 -10 L-glutamate EX_cpd00023_e0 0 -0.0283302 0 D-glucose EX_cpd00027_e0 -0.61972444 -0.04098397 0 Mn2 EX_cpd00030_e0 -15 -15 -10 Glycine EX_cpd00033_e0 -0.0068175 -0.00693094 0 Zn2 EX_cpd00034_e0 -15 -15 -10 L-alanine EX_cpd00035_e0 -0.02780553 -0.00823049 0 Succinate EX_cpd00036_e0 -0.0056245 -0.12240603 0 L-lysine EX_cpd00039_e0 0 -10 0 L-aspartate EX_cpd00041_e0 0 -0.03205557 0 Sulfate EX_cpd00048_e0 -15 -15 -10 L-arginine EX_cpd00051_e0 -0.0068175 -0.00948672 0 L-serine EX_cpd00054_e0 0 -0.01004986 0 Cu2+ EX_cpd00058_e0 -15 -15 -10 Ca2+ EX_cpd00063_e0 -15 -100 -10 L-ornithine EX_cpd00064_e0 -0.0068175 -0.00831712 0 H+ EX_cpd00067_e0 -15 -15 -10 L-tyrosine EX_cpd00069_e0 -0.0068175 -0.00233919 0 Sucrose EX_cpd00076_e0 0 -0.02049199 0 L-cysteine EX_cpd00084_e0 -0.0068175 0 0 Cl- EX_cpd00099_e0 -15 -15 -10 Glycerol EX_cpd00100_e0 0 0 -10 Biotin EX_cpd00104_e0 -15 -15 0 D-ribose EX_cpd00105_e0 -0.01862144 0 0 L-leucine EX_cpd00107_e0 -0.03596182 -0.00303228 0 D-galactose EX_cpd00108_e0 -0.25290619 -0.18317325 0 L-histidine EX_cpd00119_e0 -0.0068175 -0.00506825 0 L-proline EX_cpd00129_e0 -0.01102953 0 0 L-malate EX_cpd00130_e0 -0.03649016 -0.79413596 0 D-mannose EX_cpd00138_e0 -0.2540567 -0.05436649 0 Co2 EX_cpd00149_e0 -
Expanding Metabolism for Total Biosynthesis of the Nonnatural Amino Acid L-Homoalanine
Expanding metabolism for total biosynthesis of the nonnatural amino acid L-homoalanine Kechun Zhanga, Han Lib, Kwang Myung Choa, and James C. Liaoa,b,c,1 aDepartment of Chemical and Biomolecular Engineering, University of California, Los Angeles, CA 90095; bMolecular Biology Institute, University of California, Los Angeles, CA 90095; and cInstitute for Genomics and Proteomics, University of California, Los Angeles, CA 90095 Edited by James A. Wells, University of California, San Francisco, CA, and approved January 25, 2010 (received for review November 8, 2009) The dramatic increase in healthcare cost has become a significant L-glutamate, L-lysine, and L-threonine are produced more than burden to the world. Many patients are denied the accessibility of 2 million tons annually (5). Unlike natural amino acids, total bio- medication because of the high price of drugs. Total biosynthesis of synthesis of nonnatural amino acids from simple sugars encoun- chiral drug intermediates is an environmentally friendly approach ters significant technical challenges: First, artificial metabolic that helps provide more affordable pharmaceuticals. Here we have pathways have to be designed to expand the metabolic capability expanded the natural metabolic capability to biosynthesize a non- of cells (8); second, optimizing the designed unique metabolic natural amino acid L-homoalanine, which is a chiral precursor of pathways requires extensive protein evolution (9); third, meta- levetiracetam, brivaracetam, and ethambutol. We developed a se- bolic flux should be driven to the production of the target com- lection strategy and altered the substrate specificity of ammonium- pounds (10). We have successfully addressed these problems and assimilating enzyme glutamate dehydrogenase. -
O O2 Enzymes Available from Sigma Enzymes Available from Sigma
COO 2.7.1.15 Ribokinase OXIDOREDUCTASES CONH2 COO 2.7.1.16 Ribulokinase 1.1.1.1 Alcohol dehydrogenase BLOOD GROUP + O O + O O 1.1.1.3 Homoserine dehydrogenase HYALURONIC ACID DERMATAN ALGINATES O-ANTIGENS STARCH GLYCOGEN CH COO N COO 2.7.1.17 Xylulokinase P GLYCOPROTEINS SUBSTANCES 2 OH N + COO 1.1.1.8 Glycerol-3-phosphate dehydrogenase Ribose -O - P - O - P - O- Adenosine(P) Ribose - O - P - O - P - O -Adenosine NICOTINATE 2.7.1.19 Phosphoribulokinase GANGLIOSIDES PEPTIDO- CH OH CH OH N 1 + COO 1.1.1.9 D-Xylulose reductase 2 2 NH .2.1 2.7.1.24 Dephospho-CoA kinase O CHITIN CHONDROITIN PECTIN INULIN CELLULOSE O O NH O O O O Ribose- P 2.4 N N RP 1.1.1.10 l-Xylulose reductase MUCINS GLYCAN 6.3.5.1 2.7.7.18 2.7.1.25 Adenylylsulfate kinase CH2OH HO Indoleacetate Indoxyl + 1.1.1.14 l-Iditol dehydrogenase L O O O Desamino-NAD Nicotinate- Quinolinate- A 2.7.1.28 Triokinase O O 1.1.1.132 HO (Auxin) NAD(P) 6.3.1.5 2.4.2.19 1.1.1.19 Glucuronate reductase CHOH - 2.4.1.68 CH3 OH OH OH nucleotide 2.7.1.30 Glycerol kinase Y - COO nucleotide 2.7.1.31 Glycerate kinase 1.1.1.21 Aldehyde reductase AcNH CHOH COO 6.3.2.7-10 2.4.1.69 O 1.2.3.7 2.4.2.19 R OPPT OH OH + 1.1.1.22 UDPglucose dehydrogenase 2.4.99.7 HO O OPPU HO 2.7.1.32 Choline kinase S CH2OH 6.3.2.13 OH OPPU CH HO CH2CH(NH3)COO HO CH CH NH HO CH2CH2NHCOCH3 CH O CH CH NHCOCH COO 1.1.1.23 Histidinol dehydrogenase OPC 2.4.1.17 3 2.4.1.29 CH CHO 2 2 2 3 2 2 3 O 2.7.1.33 Pantothenate kinase CH3CH NHAC OH OH OH LACTOSE 2 COO 1.1.1.25 Shikimate dehydrogenase A HO HO OPPG CH OH 2.7.1.34 Pantetheine kinase UDP- TDP-Rhamnose 2 NH NH NH NH N M 2.7.1.36 Mevalonate kinase 1.1.1.27 Lactate dehydrogenase HO COO- GDP- 2.4.1.21 O NH NH 4.1.1.28 2.3.1.5 2.1.1.4 1.1.1.29 Glycerate dehydrogenase C UDP-N-Ac-Muramate Iduronate OH 2.4.1.1 2.4.1.11 HO 5-Hydroxy- 5-Hydroxytryptamine N-Acetyl-serotonin N-Acetyl-5-O-methyl-serotonin Quinolinate 2.7.1.39 Homoserine kinase Mannuronate CH3 etc. -
Affy ID Fold Change P-Value Realtive Change Annotation Derivation
Supplemental Table 2: Gene changes at 60 minutes in adherent versus non-adherent samples Affy_ID Fold change p-value Realtive_change Annotation Derivation of annotation A4052_s_at 4.313332633 0.024012538 Increase_in_Adherent trpE; anthranilate synthase component I (EC:4.1.3.27); K01657 anthranilate synthase component I [EC:4.1.3.27] ecw:EcE24377A_1463 V1799_s_at 3.972247924 0.033307448 Increase_in_Adherent hscB; co-chaperone HscB ; K04082 molecular chaperone HscB sdy:SDY_2723 V0231_s_at 3.95580421 0.000284517 Increase_in_Adherent ssb; single-stranded DNA-binding protein ; K03111 single-strand DNA-binding protein sdy:SDY_4508 SB5_0275_s_at 3.893614168 1.07E-05 Increase_in_Adherent hypothetical protein ; K02040 phosphate transport system substrate-binding protein sbo:SBO_3421 K3820_x_at 3.763692747 7.60E-05 Increase_in_Adherent putative holin protein eoj:ECO26_3693 O2ColV121_at 3.647435418 2.22E-05 Increase_in_Adherent NO_KEGG_DATA NO_KEGG_DATA SD1_2456_x_at 3.566304619 0.015156769 Increase_in_Adherent putative fructose-like phosphotransferase EIIB subunit 3 ; K11202 PTS system, fructose-specific IIB-like component [EC:2.7.1.69] sbc:SbBS512_E4440 SDY_4180_x_at 3.540687986 4.18E-05 Increase_in_Adherent NO_KEGG_DATA NO_KEGG_DATA D1328_s_at 3.533442346 0.004222188 Increase_in_Adherent NO_KEGG_DATA NO_KEGG_DATA A0552_s_at 3.51323466 0.019815913 Increase_in_Adherent hscB; co-chaperone HscB ; K04082 molecular chaperone HscB sdy:SDY_2723 b3350_s_at 3.455476842 0.000165312 Increase_in_Adherent NO_KEGG_DATA NO_KEGG_DATA V0624_x_at 3.388319399 0.000108353 -
Mycobacterium Tuberculosis to Eradicate Persistent Infection
Research Collection Journal Article Derailing the aspartate pathway of Mycobacterium tuberculosis to eradicate persistent infection Author(s): Hasenoehrl, Erik J.; Sajorda, Dannah R.; Berney-Meyer, Linda; Johnson, Samantha; Tufariello, JoAnn M.; Fuhrer, Tobias; Cook, Gregory M.; Jacobs, William R., Jr.; Berney, Michael Publication Date: 2019 Permanent Link: https://doi.org/10.3929/ethz-b-000367423 Originally published in: Nature Communications 10, http://doi.org/10.1038/s41467-019-12224-3 Rights / License: Creative Commons Attribution 4.0 International This page was generated automatically upon download from the ETH Zurich Research Collection. For more information please consult the Terms of use. ETH Library ARTICLE https://doi.org/10.1038/s41467-019-12224-3 OPEN Derailing the aspartate pathway of Mycobacterium tuberculosis to eradicate persistent infection Erik J. Hasenoehrl1, Dannah Rae Sajorda1, Linda Berney-Meyer1, Samantha Johnson1, JoAnn M. Tufariello1,6, Tobias Fuhrer 2, Gregory M. Cook3,4, William R. Jacobs Jr.1,5 & Michael Berney 1 A major constraint for developing new anti-tuberculosis drugs is the limited number of validated targets that allow eradication of persistent infections. Here, we uncover a vulner- 1234567890():,; able component of Mycobacterium tuberculosis (Mtb) persistence metabolism, the aspartate pathway. Rapid death of threonine and homoserine auxotrophs points to a distinct sus- ceptibility of Mtb to inhibition of this pathway. Combinatorial metabolomic and tran- scriptomic analysis reveals that inability to produce threonine leads to deregulation of aspartate kinase, causing flux imbalance and lysine and DAP accumulation. Mtb’s adaptive response to this metabolic stress involves a relief valve-like mechanism combining lysine export and catabolism via aminoadipate. -
Supplemental Table S1: Comparison of the Deleted Genes in the Genome-Reduced Strains
Supplemental Table S1: Comparison of the deleted genes in the genome-reduced strains Legend 1 Locus tag according to the reference genome sequence of B. subtilis 168 (NC_000964) Genes highlighted in blue have been deleted from the respective strains Genes highlighted in green have been inserted into the indicated strain, they are present in all following strains Regions highlighted in red could not be deleted as a unit Regions highlighted in orange were not deleted in the genome-reduced strains since their deletion resulted in severe growth defects Gene BSU_number 1 Function ∆6 IIG-Bs27-47-24 PG10 PS38 dnaA BSU00010 replication initiation protein dnaN BSU00020 DNA polymerase III (beta subunit), beta clamp yaaA BSU00030 unknown recF BSU00040 repair, recombination remB BSU00050 involved in the activation of biofilm matrix biosynthetic operons gyrB BSU00060 DNA-Gyrase (subunit B) gyrA BSU00070 DNA-Gyrase (subunit A) rrnO-16S- trnO-Ala- trnO-Ile- rrnO-23S- rrnO-5S yaaC BSU00080 unknown guaB BSU00090 IMP dehydrogenase dacA BSU00100 penicillin-binding protein 5*, D-alanyl-D-alanine carboxypeptidase pdxS BSU00110 pyridoxal-5'-phosphate synthase (synthase domain) pdxT BSU00120 pyridoxal-5'-phosphate synthase (glutaminase domain) serS BSU00130 seryl-tRNA-synthetase trnSL-Ser1 dck BSU00140 deoxyadenosin/deoxycytidine kinase dgk BSU00150 deoxyguanosine kinase yaaH BSU00160 general stress protein, survival of ethanol stress, SafA-dependent spore coat yaaI BSU00170 general stress protein, similar to isochorismatase yaaJ BSU00180 tRNA specific adenosine