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Homology modeling of the CheW coupling protein of the chemotaxis signaling complex.

ABSTRACT: Homology models of the E. coli and T. maritima chemotaxis protein CheW were constructed to assess the quality of structural predictions and their applicability in chemotaxis research: i) a model of E. coli CheW was constructed using the T. maritima CheW NMR structure as a template, and ii) a model of T. maritima CheW was constructed using the E. coli CheW NMR structure as a template. The conformational space accessible to the homology models and to the NMR structures was investigated using molecular dynamics and Monte Carlo simulations. The results show that even though static homology models of CheW may be partially structurally different from their corresponding experimentally determined structures, the conformational space they can access through their dynamic variations can be similar, for specific regions of the protein, to that of the experimental NMR structures. When CheW homology models are allowed to explore their local accessible conformational space, modeling can provide a rational path to predicting CheW interactions with the MCP and CheA proteins of the chemotaxis complex. Homology models of CheW (and potentially, of other chemotaxis proteins) should be seen as snapshots of an otherwise larger ensemble of accessible conformational space.

SUBMITTER: Cashman DJ

PROVIDER: S-EPMC3737408 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Homology modeling of the CheW coupling protein of the chemotaxis signaling complex.

Cashman Derek J DJ Ortega Davi R DR Zhulin Igor B IB Baudry Jerome J

PloS one 20130807 8

Homology models of the E. coli and T. maritima chemotaxis protein CheW were constructed to assess the quality of structural predictions and their applicability in chemotaxis research: i) a model of E. coli CheW was constructed using the T. maritima CheW NMR structure as a template, and ii) a model of T. maritima CheW was constructed using the E. coli CheW NMR structure as a template. The conformational space accessible to the homology models and to the NMR structures was investigated using molec ...[more]

PMID: 23950985

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Project description:The signaling apparatus that controls bacterial chemotaxis is composed of a core complex containing chemoreceptors, the histidine autokinase CheA, and the coupling protein CheW. Site-specific spin labeling and pulsed dipolar ESR spectroscopy (PDS) have been applied to investigate the structure of a soluble ternary complex formed by Thermotoga maritima CheA (TmCheA), CheW, and receptor signaling domains. Thirty-five symmetric spin-label sites (SLSs) were engineered into the five domains of the CheA dimer and CheW to provide distance restraints within the CheA:CheW complex in the absence and presence of a soluble receptor that inhibits kinase activity (Tm14). Additional PDS restraints among spin-labeled CheA, CheW, and an engineered single-chain receptor labeled at six different sites allow docking of the receptor structure relative to the CheA:CheW complex. Disulfide cross-linking between selectively incorporated Cys residues finds two pairs of positions that provide further constraints within the ternary complex: one involving Tm14 and CheW and another involving Tm14 and CheA. The derived structure of the ternary complex indicates a primary site of interaction between CheW and Tm14 that agrees well with previous biochemical and genetic data for transmembrane chemoreceptors. The PDS distance distributions are most consistent with only one CheW directly engaging one dimeric Tm14. The CheA dimerization domain (P3) aligns roughly antiparallel to the receptor-conserved signaling tip but does not interact strongly with it. The angle of the receptor axis with respect to P3 and the CheW-binding P5 domains is bound by two limits differing by approximately 20 degrees . In one limit, Tm14 aligns roughly along P3 and may interact to some extent with the hinge region near the P3 hairpin loop. In the other limit, Tm14 tilts to interact with the P5 domain of the opposite subunit in an interface that mimics that observed with the P5 homologue CheW. The time domain ESR data can be simulated from the model only if orientational variability is introduced for the P5 and, especially, P3 domains. The Tm14 tip also binds beside one of the CheA kinase domains (P4); however, in both bound and unbound states, P4 samples a broad range of distributions that are only minimally affected by Tm14 binding. The CheA P1 domains that contain the substrate histidine are also broadly distributed in space under all conditions. In the context of the hexagonal lattice formed by trimeric transmembrane chemoreceptors, the PDS structure is best accommodated with the P3 domain in the center of a honeycomb edge.

Project description:Salmonella enterica is the most frequently reported cause of foodborne illness. As in other microorganisms, chemotaxis affords key physiological benefits, including enhanced access to growth substrates, but also plays an important role in infection and disease. Chemoreceptor signaling core complexes, consisting of CheA, CheW and methyl-accepting chemotaxis proteins (MCPs), modulate the switching of bacterial flagella rotation that drives cell motility. These complexes, through the formation of heterohexameric rings composed of CheA and CheW, form large clusters at the cell poles. RecA plays a key role in polar cluster formation, impairing the assembly when the SOS response is activated. In this study, we determined that RecA protein interacts with both CheW and CheA. The binding of these proteins to RecA is needed for wild-type polar cluster formation. In silico models showed that one RecA molecule, attached to one signaling unit, fits within a CheA-CheW ring without interfering with the complex formation or array assembly. Activation of the SOS response is followed by an increase in RecA, which rises up the number of signaling complexes associated with this protein. This suggests the presence of allosteric inhibition in the CheA-CheW interaction and thus of heterohexameric ring formation, impairing the array assembly. STED imaging demonstrated that all core unit components (CheA, CheW, and MPCs) have the same subcellular location as RecA. Activation of the SOS response promotes the RecA distribution along the cell instead of being at the cell poles. CheA- and CheW- RecA interactions are also crucial for chemotaxis, which is maintained when the SOS response is induced and the signaling units are dispersed. Our results provide new molecular-level insights into the function of RecA in chemoreceptor clustering and chemotaxis determining that the impaired chemoreceptor clustering not only inhibits swarming but also modulates chemotaxis in SOS-induced cells, thereby modifying bacterial motility in the presence of DNA-damaging compounds, such as antibiotics.

Dataset Information

Homology modeling of the CheW coupling protein of the chemotaxis signaling complex.

Publications

Homology modeling of the CheW coupling protein of the chemotaxis signaling complex.

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