Article

Solution structure of the NEAT (NEAr Transporter) domain from IsdH/HarA: the human hemoglobin receptor in Staphylococcus aureus.

Department of Chemistry and Biochemistry, UCLA-DOE Institute for Genomics and Proteomics, and the Molecular Biology Institute, University of California-Los Angeles, 611 Charles E. Young Drive, Los Angeles, CA 90095-1570, USA.
Journal of Molecular Biology (Impact Factor: 3.96). 08/2006; 360(2):435-47. DOI: 10.1016/j.jmb.2006.05.019
Source: PubMed

ABSTRACT During infections the pathogen Staphylococcus aureus procures the essential nutrient iron from its host using iron-regulated surface determinant (Isd) proteins, which scavenge heme bound iron from host hemoproteins. Four Isd proteins are displayed in the cell wall, where they function as receptors for host proteins and heme. Each of the receptors contains one or more copies of a recently discovered domain called NEAT (NEAr Transporter) that has been shown to mediate protein binding. Here we report the three-dimensional solution structure of the NEAT domain from the IsdH/HarA protein, which is the hemoglobin receptor in the Isd system. This is the first structure of a NEAT domain and reveals that they adopt a beta sandwich fold that consists of two five-stranded antiparallel beta sheets. Although unrelated at the primary sequence level, our results indicate that NEAT domains belong to the immunoglobulin superfamily. Binding studies indicate that two IsdH/HarA NEAT domains bind a single molecule of methemoglobin, while the distantly related NEAT domain from the S. aureus IsdC protein binds only heme. A comparison of their primary sequences in light of the new structure is used to predict the hemoglobin and heme binding surfaces on NEAT domains.

0 Followers
 · 
64 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Haem is the major iron source for bacteria that develop in higher organisms. In these hosts, bacteria have to cope with nutritional immunity imposed by the host, since haem and iron are tightly bound to carrier and storage proteins. Siderophores were the first recognized fighters in the battle for iron between bacteria and host. They are non-proteinaceus organic molecules having an extremely high affinity for Fe(3+) and able to extract it from host proteins. Haemophores, that display functional analogy with siderophores, were more recently discovered. They are a class of secreted proteins with a high affinity for haem; they are able to extract haem from host haemoproteins and deliver it to specific receptors that internalize haem. In the past few years, a wealth of data has accumulated on haem acquisition systems that are dependent on surface exposed/secreted bacterial proteins. They promote haem transfer from its initial source (in most cases, a eukaryotic haem binding protein) to the transporter that carries out the membrane crossing step. Here we review recent discoveries in this field, with particular emphasis on similar and dissimilar mechanisms in haemophores and siderophores, from the initial host source to the binding protein/receptor at the cell surface.
    Molecular Microbiology 06/2012; 85(4):618-31. DOI:10.1111/j.1365-2958.2012.08136.x · 5.03 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: A growing body of evidence suggests that surface or secreted proteins with NEAr Transporter (NEAT) domains play a central role in haem acquisition and trafficking across the cell envelope of Gram-positive bacteria. Group A streptococcus (GAS), a β-haemolytic human pathogen, expresses a NEAT protein, Shr, which binds several haemoproteins and extracellular matrix (ECM) components. Shr is a complex, membrane-anchored protein, with a unique N-terminal domain (NTD) and two NEAT domains separated by a central leucine-rich repeat region. In this study we have carried out an analysis of the functional domains in Shr. We show that Shr obtains haem in solution and furthermore reduces the haem iron; this is the first report of haem reduction by a NEAT protein. More specifically, we demonstrate that both of the constituent NEAT domains of Shr are responsible for binding haem, although they are missing a critical tyrosine residue found in the ligand-binding pocket of other haem-binding NEAT domains. Further investigations show that a previously undescribed region within the Shr NTD interacts with methaemoglobin. Shr NEAT domains, however, do not contribute significantly to the binding of methaemoglobin but mediate binding to the ECM components fibronectin and laminin. A protein fragment containing the NTD plus the first NEAT domain was found to be sufficient to sequester haem directly from methaemoglobin. Correlating these in vitro findings to in vivo biological function, mutants analysis establishes the role of Shr in GAS growth with methaemoglobin as a sole source of iron, and indicates that at least one NEAT domain is necessary for the utilization of methaemoglobin. We suggest that Shr is the prototype of a new group of NEAT composite proteins involved in haem uptake found in pyogenic streptococci and Clostridium novyi.
    Molecular Microbiology 11/2010; 78(3):739-56. DOI:10.1111/j.1365-2958.2010.07367.x · 5.03 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Three iron-regulated surface determinant (Isd) proteins, containing NEAr Transporter (NEAT) domains (GBAA4789-7), constitute part of an eight-member Bacillus anthracis operon. GBAA4789 (IsdC), previously characterized by others as a haem-binding protein, and two novel Isd proteins characterized in this study, GBAA4788 (IsdJ) and GBAA4787 (IsdK) proteins, can be translated from two alternative overlapping transcriptional units. The three NEAT-containing Isd proteins are shown to be expressed in vivo during B. anthracis infection. Expression in vitro is regulated by iron ions independent of the virulence plasmids pXO1 and pXO2, yet their presence affects the range of response to iron ion concentration. The expression of IsdC, J and K is strongly repressed under high CO(2) tension, conditions that are optimal for B. anthracis toxin and capsule expression, suggesting that these Isd proteins are elements of a B. anthracis'air-regulon'. Deletion mutants of isdC, isdK or the entire isdCJK locus are as virulent and pathogenic to guinea pigs as the fully virulent wild-type Vollum strain. The isdC-deleted mutant is defective in sequestration of haemin, consistent with previous biochemical observations, while the DeltaisdK mutant is defective in haemoglobin uptake. Studies with recombinant IsdK demonstrate specific binding to haemoglobin.
    Molecular Microbiology 10/2008; 70(4):983-99. DOI:10.1111/j.1365-2958.2008.06460.x · 5.03 Impact Factor