title: Spectroscopic Determination of Distinct Heme Ligands in Outer-Membrane Receptors PhuR and HasR of Pseudomonas aeruginosa.
abstract: Pseudomonas aeruginosa PAO1 encodes two outer membrane receptors, PhuR (Pseudomonas heme uptake) and HasR (heme assimilation system). The HasR receptor acquires heme through interaction with a secreted hemophore, HasAp. The non-hemophore-dependent PhuR is encoded along with proteins required for heme translocation into the cytoplasm. Herein, we report the isolation and characterization of the HasR and PhuR receptors. Absorption and MCD spectroscopy confirmed that, similar to other Gram-negative OM receptors, HasR coordinates heme through the conserved N-terminal plug His-221 and His-624 of the surface-exposed FRAP-loop. In contrast, PhuR showed distinct absorption and MCD spectra consistent with coordination through a Tyr residue. Sequence alignment of PhuR with all known Gram-negative OM heme receptors revealed a lack of a conserved His within the FRAP loop but two Tyr residues at positions 519 and 529. Site-directed mutagenesis and spectroscopic characterization confirmed Tyr-519 and the N-terminal plug His-124 provide the heme ligands in PhuR. We propose that PhuR and HasR represent nonredundant heme receptors capable of sensing and accessing heme across a wide range of physiological conditions on colonization and infection of the host.
authors: †Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland, Baltimore, Maryland 21201, United States.
authors: Aaron D
authors: AD
authors: Smith
authors: ‡Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208, United States.
authors: Anuja R
authors: AR
authors: Modi
authors: ‡Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208, United States.
authors: Shengfang
authors: S
authors: Sun
authors: ‡Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208, United States.
authors: John H
authors: JH
authors: Dawson
authors: †Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland, Baltimore, Maryland 21201, United States.
authors: Angela
authors: A
authors: Wilks
mesh_terms: D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006418:Heme / Q000737:chemistry* / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D011550:Pseudomonas aeruginosa / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D006418:Heme