title: Hydrogen peroxide-mediated conversion of coproheme to heme b by HemQ-lessons from the first crystal structure and kinetic studies.
abstract: Heme biosynthesis in Gram-positive bacteria follows a recently described coproporphyrin-dependent pathway with HemQ catalyzing the decarboxylation of coproheme to heme b. Here we present the first crystal structure of a HemQ (homopentameric coproheme-HemQ from Listeria monocytogenes) at 1.69 Å resolution and the conversion of coproheme to heme b followed by UV-vis and resonance Raman spectroscopy as well as mass spectrometry. The ferric five-coordinated coproheme iron of HemQ is weakly bound by a neutral proximal histidine H174. In the crystal structure of the resting state, the distal Q187 (conserved in Firmicutes HemQ) is H-bonded with propionate p2 and the hydrophobic distal cavity lacks solvent water molecules. Two H2O2molecules are shown to be necessary for decarboxylation of the propionates p2 and p4, thereby forming the corresponding vinyl groups of heme b. The overall reaction is relatively slow (kcat/KM= 1.8 × 102m-1·s-1at pH 7.0) and occurs in a stepwise manner with a three-propionate intermediate. We present the noncovalent interactions between coproheme and the protein and propose a two-step reaction mechanism. Furthermore, the structure of coproheme-HemQ is compared to that of the phylogenetically related heme b-containing chlorite dismutases.
  
  
  
  Structural data are available in the PDB under the accession number 5LOQ.
authors.affiliation: Department for Structural and Computational Biology, Max F. Perutz Laboratories, University of Vienna, Austria.
authors.forename: Stefan
authors.initials: S
authors.lastname: Hofbauer
authors.affiliation: Department for Structural and Computational Biology, Max F. Perutz Laboratories, University of Vienna, Austria.
authors.forename: Georg
authors.initials: G
authors.lastname: Mlynek
authors.affiliation: Dipartimento di Chimica 'Ugo Schiff', Università di Firenze, Sesto Fiorentino (FI), Italy.
authors.forename: Lisa
authors.initials: L
authors.lastname: Milazzo
authors.affiliation: Department for Structural and Computational Biology, Max F. Perutz Laboratories, University of Vienna, Austria.|Division of Biochemistry, Department of Chemistry, BOKU - University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Dominic
authors.initials: D
authors.lastname: Pühringer
authors.affiliation: Division of Biochemistry, Department of Chemistry, BOKU - University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Daniel
authors.initials: D
authors.lastname: Maresch
authors.affiliation: Division of Biochemistry, Department of Chemistry, BOKU - University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Irene
authors.initials: I
authors.lastname: Schaffner
authors.affiliation: Division of Biochemistry, Department of Chemistry, BOKU - University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Paul G
authors.initials: PG
authors.lastname: Furtmüller
authors.affiliation: Dipartimento di Chimica 'Ugo Schiff', Università di Firenze, Sesto Fiorentino (FI), Italy.
authors.forename: Giulietta
authors.initials: G
authors.lastname: Smulevich
authors.affiliation: Department for Structural and Computational Biology, Max F. Perutz Laboratories, University of Vienna, Austria.|Department of Biochemistry, Faculty of Chemistry and Chemical Technology, University of Ljubljana, Slovenia.
authors.forename: Kristina
authors.initials: K
authors.lastname: Djinović-Carugo
authors.affiliation: Division of Biochemistry, Department of Chemistry, BOKU - University of Natural Resources and Life Sciences, Vienna, Austria.
authors.forename: Christian
authors.initials: C
authors.lastname: Obinger
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006094:Gram-Positive Bacteria / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D006418:Heme / Q000737:chemistry / Q000378:metabolism*; D006420:Hemeproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006427:Hemin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D006860:Hydrogen Bonding; D006861:Hydrogen Peroxide / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008089:Listeria monocytogenes / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D013058:Mass Spectrometry; D008958:Models, Molecular; D011422:Propionates / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D000072417:Protein Domains; D017386:Sequence Homology, Amino Acid; D013053:Spectrophotometry; D013059:Spectrum Analysis, Raman
chemical_list: D001426:Bacterial Proteins; D006420:Hemeproteins; D011422:Propionates; C561849:coprohemin; D006418:Heme; D006427:Hemin; D006861:Hydrogen Peroxide
keywords: Gram-positive pathogens; HemQ; coproheme; heme b; heme biosynthesis