38152937
Structural Mechanisms of QacR Induction and Multidrug Recognition
The Staphylococcus aureus multidrug binding protein QacR represses transcription of the qacA multidrug transporter gene and is induced by structurally diverse cationic lipophilic drugs. Here, we report the crystal structures of six QacR-drug complexes. Compared to the DNA bound structure, drug binding elicits a coil-to-helix transition that causes induction and creates an expansive multidrug-binding pocket, containing four glutamates and multiple aromatic and polar residues. These structures indicate the presence of separate but linked drug-binding sites within a single protein. This multisite drug-binding mechanism is consonant with studies on multidrug resistance transporters.
19913942
Crystal Structures of QacR-Diamidine Complexes Reveal Additional Multidrug-binding Modes and a Novel Mechanism of Drug Charge Neutralization*
The Staphylococcus aureus multidrug-binding protein QacR represses transcription of the plasmid-encoded membrane protein QacA, a multidrug efflux transporter. QacR is induced by multiple structurally dissimilar monovalent and bivalent cationic lipophilic compounds, many of which are effluxed from the cell by QacA via the proton motive force. The multidrug-binding pocket of QacR has been shown to be quite extensive and features several glutamates and multiple aromatic residues. To date, the structure of only one QacR-bivalent cationic drug complex (that of QacR bound to dequalinium) has been determined, and how other longer or shorter bivalent cationic compounds bind is unknown. Here we report the crystal structures of QacR bound to two cytotoxic bivalent diamidines, pentamidine and hexamidine. These compounds are structurally similar, differing by only one methylene carbon in the alkyl chain linker. However, this small difference results in very dissimilar binding modes. Similar to dequalinium, hexamidine spans the multidrug-binding pocket, and its positively charged benzamidine groups are neutralized by residues Glu-57 and Glu-120. Pentamidine binds QacR in a novel fashion whereby one of its benzamidine groups interacts with residue Glu-63, and the other is neutralized by carbonyl and side chain oxygen atoms. Thus, these structures demonstrate that a formal negative charge is not a prerequisite for binding positively charged drugs and underscore the versatility of the QacR and, likely, all multidrug-binding pockets.
4431121
Crystal structures of SarA, a pleiotropic regulator of virulence genes in S. aureus
Staphylococcus aureus is a major human pathogen, the potency of which can be attributed to the regulated expression of an impressive array of virulence determinants. A key pleiotropic transcriptional regulator of these virulence factors is SarA, which is encoded by the sar (staphylococcal accessory regulator) locus. SarA was characterized initially as an activator of a second virulence regulatory locus, agr, through its interaction with a series of heptad repeats (AGTTAAG) within the agr promoter. Subsequent DNA-binding studies have revealed that SarA binds readily to multiple AT-rich sequences of variable lengths. Here we describe the crystal structure of SarA and a SarA–DNA complex at resolutions of 2.50 Å and 2.95 Å, respectively. SarA has a fold consisting of a four-helix core region and ‘inducible regions’ comprising a β-hairpin and a carboxy-terminal loop. On binding DNA, the inducible regions undergo marked conformational changes, becoming part of extended and distorted α-helices, which encase the DNA. SarA recognizes an AT-rich site in which the DNA is highly overwound and adopts a D-DNA-like conformation by indirect readout. These structures thus provide insight into SarA-mediated transcription regulation.
29559148
Structural basis for the recognition of a bisphosphorylated MAP kinase peptide by human VHR protein Phosphatase.
Human VHR (vaccinia H1 related phosphatase) is a member of the dual-specificity phosphatases (DSPs) that often act on bisphosphorylated protein substrates. Unlike most DSPs, VHR displays a strong preference for dephosphorylating phosphotyrosine residues over phosphothreonine residues. Here we describe the 2.75 A crystal structure of the C124S inactive VHR mutant in complex with a bisphosphorylated peptide corresponding to the MAP kinase activation lip. This structure and subsequent biochemical studies revealed the basis for the strong preference for hydrolyzing phosphotyrosine within bisphosphorylated substrates containing -pTXpY-. In the structure, the two phospho residues are oriented into distinct pockets; the phosphotyrosine is bound in the exposed yet deep active site cleft while the phosphothreonine is loosely tethered into a nearby basic pocket containing Arg(158). As this structure is the first substrate-enzyme complex reported for the DSP family of enzymes, these results provide the first glimpse into how DSPs bind their protein substrates.
5803090
Structural basis for cooperative DNA binding by two dimers of the multidrug-binding protein QacR.
The Staphylococcus aureus multidrug-binding protein QacR represses transcription of the qacA multidrug transporter gene and is induced by multiple structurally dissimilar drugs. QacR is a member of the TetR/CamR family of transcriptional regulators, which share highly homologous N-terminal DNA-binding domains connected to seemingly non-homologous ligand-binding domains. Unlike other TetR members, which bind approximately 15 bp operators, QacR recognizes an unusually long 28 bp operator, IR1, which it appears to bind cooperatively. To elucidate the DNA-binding mechanism of QacR, we determined the 2.90 A resolution crystal structure of a QacR-IR1 complex. Strikingly, our data reveal that the DNA recognition mode of QacR is distinct from TetR and involves the binding of a pair of QacR dimers. In this unique binding mode, recognition at each IR1 half-site is mediated by a complement of DNA contacts made by two helix-turn-helix motifs. The inferred cooperativity does not arise from cross-dimer protein-protein contacts, but from the global undertwisting and major groove widening elicited by the binding of two QacR dimers.
22785877
Structures of the pleiotropic translational regulator Hfq and an Hfq-RNA complex: a bacterial Sm-like protein.
In prokaryotes, Hfq regulates translation by modulating the structure of numerous RNA molecules by binding preferentially to A/U-rich sequences. To elucidate the mechanisms of target recognition and translation regulation by Hfq, we determined the crystal structures of the Staphylococcus aureus Hfq and an Hfq-RNA complex to 1.55 and 2.71 A resolution, respectively. The structures reveal that Hfq possesses the Sm-fold previously observed only in eukaryotes and archaea. However, unlike these heptameric Sm proteins, Hfq forms a homo-hexameric ring. The Hfq-RNA structure reveals that the single-stranded hepta-oligoribonucleotide binds in a circular conformation around a central basic cleft, whereby Tyr42 residues from adjacent subunits stack with six of the bases, and Gln8, outside the Sm motif, provides key protein-base contacts. Such binding suggests a mechanism for Hfq function.
118275903
CP Tests and the Weak Dipole Moment in τ-Pair Production at LEP
Tests of CP in variance in the reaction e +e− →τ+τ− on the Z 0 peak are reviewed. Averaging the present LEP measurements, upper limits on the real and imaginary parts of the weak dipole moment of the τ lepton of ∣Re(d τ w (M Z 2 ))∣ < 3.3 • 10−18 ecm and ∣Im(d τ w (M Z 2 ))∣ < 9.8 • 10−18 ecm with 95% c.l. are obtained.
117936692
TPC with GEM readout for TESLA
A Time Projection Chamber (TPC) is proposed as the main tracking device for the detector at the future e+e− linear accelerator now studied in Europe. The performance requirements and the basic design using a Gas Electron Multiplier (GEM) as the readout device are described. Some preliminary R&D results are discussed.
6747128
Prospects for the measurement of the structure of the coupling of a Higgs boson to weak gauge bosons in weak boson fusion with the ATLAS detector
The prospects for the measurement of the tensor structure of the vertex between a standard model Higgs boson and two weak gauge bosons using the distribution of the azimuthal angles between the two tagging jets in the weak boson fusion channel are studied in a Monte Carlo analysis using the fast simulation of the ATLAS detector. The decay channels H→τ+τ-→ll+4ν, H→τ+τ-→lh+3ν at mH=120 GeV and H→W+W-→llνν at mH=160 GeV are used in the analysis. For a standard model Higgs boson it is found that purely anomalous couplings are expected to be excluded at a confidence level corresponding to 2σ or more at mH=120 GeV and more than 5σ at mH=160 GeV from 30 fb-1 of data. With a value of 1 roughly reproducing the standard model cross section for a purely anomalous coupling, the standard deviation in a measurement of a contribution of a CP even anomalous coupling in addition to the standard model coupling is estimated to be 0.20 at mH=120 GeV and 0.09 at mH=160 GeV.
117172015
The Higgs Boson: Still Elusive After 40 Years
8372632
Investigation of the Discovery Potential for Higgs Bosons of the Minimal Supersymmetric Extension of the Standard Model (MSSM) with ATLAS
The discovery potential of the ATLAS experiment at the LHC for Higgs bosons of the MSSM is discussed. Several CP conserving and one CP violating benchmark scenario are investigated.