Novel Mechanism of Hemin Capture by Hbp2, the Hemoglobin-binding Hemophore from Listeria monocytogenes

Malmirchegini, G. Reza; Sjodt, Megan; Shnitkind, Sergey; Sawaya, M.R.; Rosinski, Justin; Newton, S. A.; Klebba, Phillip E.; Clubb, Robert T.

doi:10.1074/jbc.m114.583013

Cited by 29 publications

(32 citation statements)

References 58 publications

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“…Instead, a tyrosine on the adjacent ␤-strand (␤7) from the ␤-strand harboring the heme-binding motif (␤8) coordinates its hydroxyl group to heme-iron. Furthermore, Hbp2 N2 undergoes a major conformational change upon heme binding (90), which has not been observed for other NEAT domains, such as B. anthracis IsdX1 (56), S. aureus IsdA (69), and S. aureus IsdH (91). The authors of those studies suggest that this larger structural rearrangement of Hbp2 N2 is a product of the lack of both tyrosine residues within the heme-binding motif, since within NEAT domains with a conserved heme-binding motif, the two tyrosine residues form a noncovalent interaction that stabilizes the ␤-hairpin structural element.…”

Section: Discussionmentioning

confidence: 86%

“…Interestingly, the second NEAT domain of the proposed hemophore from Listeria monocytogenes (Hbp2 N2 ) also harbors an amino acid substitution at the second Tyr in the heme binding motif, to an alanine and not a phenylalanine, as observed for Hal N . The recently solved structure of Hbp2 N2 shows that unlike Hal N , the conserved first Tyr of the heme-binding motif does not form an axial ligand with heme-iron (90). Instead, a tyrosine on the adjacent ␤-strand (␤7) from the ␤-strand harboring the heme-binding motif (␤8) coordinates its hydroxyl group to heme-iron.…”

Section: Discussionmentioning

confidence: 99%

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Progress toward the Development of a NEAT Protein Vaccine for Anthrax Disease

et al. 2016

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Bacillus anthracis is a sporulating Gram-positive bacterium that is the causative agent of anthrax and a potential weapon of bioterrorism. The U.S.-licensed anthrax vaccine is made from an incompletely characterized culture supernatant of a nonencapsulated, toxigenic strain (anthrax vaccine absorbed [AVA]) whose primary protective component is thought to be protective antigen (PA). AVA is effective in protecting animals and elicits toxin-neutralizing antibodies in humans, but enthusiasm is dampened by its undefined composition, multishot regimen, recommended boosters, and potential for adverse reactions. Improving next-generation anthrax vaccines is important to safeguard citizens and the military. Here, we report that vaccination with recombinant forms of a conserved domain (near-iron transporter [NEAT]), common in Gram-positive pathogens, elicits protection in a murine model of B. anthracis infection. Protection was observed with both Freund's and alum adjuvants, given subcutaneously and intramuscularly, respectively, with a mixed composite of NEATs. Protection correlated with an antibody response against the NEAT domains and a decrease in the numbers of bacteria in major organs. Anti-NEAT antibodies promote opsonophagocytosis of bacilli by alveolar macrophages. To guide the development of inactive and safe NEAT antigens, we also report the crystal structure of one of the NEAT domains (Hal) and identify critical residues mediating its heme-binding and acquisition activity. These results indicate that we should consider NEAT proteins in the development of an improved antianthrax vaccine.

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Section: Discussionmentioning

confidence: 86%

Section: Discussionmentioning

confidence: 99%

Progress toward the Development of a NEAT Protein Vaccine for Anthrax Disease

et al. 2016

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“…The side chain of the second Tyr serves as a hydrogen-bond (H-bond) donor to the phenolate oxygen atom of the axial Tyr ligand, whose bent coordination geometry enforces a π-stacking interaction with the porphyrin ring. Recently, a NEAT domain, Hbp2-N2, has been described in which a tyrosine from the β-7 strand is the axial ligand, rather than one in the β-8 YXXXY motif (44). …”

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confidence: 99%

Heme Binding by Corynebacterium diphtheriae HmuT: Function and Heme Environment

et al. 2015

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The heme uptake pathway (hmu) of Corynebacterium diphtheriae utilizes multiple proteins to bind and transport heme into the cell. One of these proteins, HmuT, delivers heme to the ABC transporter HmuUV. In this study, the axial ligation of the heme in ferric HmuT is probed by examination of wild-type HmuT and a series of conserved heme pocket residue mutants, H136A, Y235A, and M292A. Characterization by UV-visible, resonance Raman, and magnetic circular dichroism spectroscopies indicate that H136 and Y235 are the axial ligands in ferric HmuT. Consistent with this assignment of axial ligands, ferric WT and H136A HmuT are difficult to reduce while Y235A reduces readily in the presence of dithionite. Raman frequencies of the FeCO distortions in WT, H136A, and Y235A HmuT–CO complexes provide further evidence for the axial ligand assignments. Additionally, the se frequencies provide insight into the nonbonding environment of the heme pocket. Ferrous Y235A and the Y235A–CO complex reveal that the imidazole of H136 exists in two forms, one neutral and one with imidazolate character, consistent with a hydrogen-bond acceptor on the H136 side of the heme. The ferric fluoride complex of Y235A reveals the presence of at least one hydrogen-bond donor on the Y235 side of the heme. Hemoglobin utilization assays showed that the axial Y235 ligand is required for heme uptake in HmuT.

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“…Both the receptors and the hemophores use the NEAT (N-terminal near-iron transporter) domains to interact with the heme moiety through a highly conserved YXXXY motif 96 . It is interesting to note that, Hbp2 (heme/hemoglobin-binding protein 2), a NEAT-domain containing hemophore in Listeria monocytogenes , can scavenge heme but its activity is dependent on a non-canonical tyrosine residue, suggesting an unprecedented mechanism of heme binding by this protein 97 . The NEAT domain has been recognized as being very important in gram-positive biology.…”

Section: The Bacterial Acquisition Of Metals From the Hostmentioning

confidence: 99%

Iron and zinc exploitation during bacterial pathogenesis

Terwilliger

Maresso

2015

Metallomics

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Ancient bacteria originated from metal-rich environments. Billions of years of evolution directed these tiny single cell creatures to exploit the versatile properties of metals in catalyzing chemical reactions and biological responses. The result is an entire metallome of proteins that use metal co-factors to facilitate key cellular process that range from the production of energy to the replication of DNA. Two key metals in this regard are iron and zinc, both abundant on Earth but not readily accessible in a human host. Instead, pathogenic bacteria must employ clever ways to acquire these metals. In this review we describe the many elegant ways these bacteria mine, regulate, and craft the use of two key metals (iron and zinc) to build a virulence arsenal that challenges even the most sophisticated immune response.

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Novel Mechanism of Hemin Capture by Hbp2, the Hemoglobin-binding Hemophore from Listeria monocytogenes

Cited by 29 publications

References 58 publications

Progress toward the Development of a NEAT Protein Vaccine for Anthrax Disease

Progress toward the Development of a NEAT Protein Vaccine for Anthrax Disease

Heme Binding by Corynebacterium diphtheriae HmuT: Function and Heme Environment

Iron and zinc exploitation during bacterial pathogenesis

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