title: Metabolic flux of extracellular heme uptake in Pseudomonas aeruginosa is driven by the iron-regulated heme oxygenase (HemO).
abstract: Heme utilization by Pseudomonas aeruginosa involves several proteins required for internalization and degradation of heme. In the following report we provide the first direct in vivo evidence for the specific degradation of extracellular heme to biliverdin (BV) by the iron-regulated HemO. Moreover, through isotopic labeling ((13)C-heme) and electrospray ionization-MS analysis we have confirmed the regioselectivity and ratio of (13)C-δ and β-BV IX (70:30) is identical in vivo to that previously observed for the purified protein. Furthermore, the (13)C-BV IXδ and BV IXβ products are effluxed from the cell by an as yet unidentified transporter. Conversion of extracellular heme to BV is dependent solely on the iron-regulated HemO as evidenced by the lack of BV production in the P. aeruginosa hemO deletion strain. Complementation of P. aeruginosa ΔhemO with a plasmid expressing either the wild type HemO or α-regioselective HemO mutant restored extracellular heme uptake and degradation. In contrast deletion of the gene encoding the cytoplasmic heme-binding protein, PhuS, homologs of which have been proposed to be heme oxygenases, did not eliminate (13)C-BV IXδ and IXβ production. In conclusion the metabolic flux of extracellular heme as a source of iron is driven by the catalytic action of HemO.
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland, Baltimore, Maryland 21201-1140, USA.
authors.forename: Kylie D
authors.initials: KD
authors.lastname: Barker
authors.forename: Katalin
authors.initials: K
authors.lastname: Barkovits
authors.forename: Angela
authors.initials: A
authors.lastname: Wilks
mesh_terms: D015153:Blotting, Western; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005816:Genetic Complementation Test; D006418:Heme / Q000378:metabolism*; D006419:Heme Oxygenase (Decyclizing) / Q000235:genetics / Q000378:metabolism*; D007501:Iron / Q000378:metabolism*; D011550:Pseudomonas aeruginosa / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D006418:Heme; D007501:Iron; D006419:Heme Oxygenase (Decyclizing)