11 research outputs found
Crystal Structure of an Invasivity-Associated Domain of SdrE in <i>S</i>. <i>aureus</i>
<div><p>The surface protein SdrE, a microbial surface components recognizing adhesive matrix molecule (MSCRAMM) family protein expressed on the surface of <i>Staphylococcus aureus</i> (<i>S</i>. <i>aureus</i>), can recognize human complement regulator Factor H and C4BP, thus making it a potentially promising vaccine candidate. In this study, SdrE<sup>278-591</sup> was found to directly affect <i>S</i>. <i>aureus</i> host cell invasion. Additionally, the crystal structure of SdrE<sup>278-591</sup> at a resolution of 1.25 Å was established, with the three-dimensional structure revealing N2-N3 domains which fold in a manner similar to an IgG fold. Furthermore, a putative ligand binding site located at a conserved charged groove formed by the interface between N2 and N3 domains was identified, with β2 suspected to occupy the ligand recognizing site and undergo a structural rearrangement to allow ligand binding. Overall, these findings have further contributed to the understanding of SdrE as a key factor for <i>S</i>. <i>aureus</i> invasivity and will enable a better understanding of bacterial infection processes.</p></div
Adherence and invasion of the ΔSdrE and ΔSdrE<sup>278-591</sup> mutants in host cell lines <i>in vitro</i>.
<p><i>S</i>. <i>aureus</i> Mu50 and its isogenic mutants <i>ΔSdrE</i> (Mu50Δ<i>SdrE</i>) and ΔSdrE<sup>278-591</sup> (Mu50Δ<i>SdrE-A</i>) were examined for adherence in HeLa (A) and 143B cells (B). These same mutants were also examined for invasivity in HeLa (C) and 143B (D) cells. Infectivity assessments were conducted for 4 h at 37°C. Wild-type <i>S</i>. <i>aureus</i> and the <i>ΔSdrE</i> (Mu50<i>ΔSdrE</i>) and <i>ΔSdrE</i><sup>278-591</sup>(Mu50<i>ΔSdrE-A</i>) mutants were generated to be devoid of SdrE in order to avoid destruction of the monolayer infection system. Scoring of the number of adherent and invasive bacterial cells indicate that adhesion and invasion are substantially reduced for <i>ΔSdrE</i> ((Mu50<i>ΔSdrE</i>) and <i>ΔSdrE</i><sup>278-591</sup> -deficient (Mu50<i>ΔSdrE-A</i>) <i>S</i>. <i>aureus</i> mutant. Results are presented as a mean ± standard deviation for at least three independent experiments. Asterisks and triangles denote values significantly different from the wild-type as determined by Student’s t-test (** P < 0.01).</p
<i>S</i>. <i>aureus</i> SdrE<sup>278-591</sup> (magenta) superimposed on its homolog <i>S</i>. <i>aureus</i> Bbp (cyan).
<p>(A) <i>S</i>. <i>aureus</i> SdrE<sup>278-591</sup> (magenta) superimposed on its homolog Bbp (substrate free, PDB code 5cf3). (B) <i>S</i>. <i>aureus</i> SdrE<sup>278-591</sup> (magenta) superimposed on its homolog Bbp (substrate, PDB code 5cfa). The predominantly green strand is the peptide ligand (substrate).</p
Data collection and refinement statistics.
<p>Data collection and refinement statistics.</p
Thermal Evaporation and Characterization of Sb<sub>2</sub>Se<sub>3</sub> Thin Film for Substrate Sb<sub>2</sub>Se<sub>3</sub>/CdS Solar Cells
Sb<sub>2</sub>Se<sub>3</sub> is a promising absorber material for
photovoltaic cells because of its optimum band gap, strong optical
absorption, simple phase and composition, and earth-abundant and nontoxic
constituents. However, this material is rarely explored for photovoltaic
application. Here we report Sb<sub>2</sub>Se<sub>3</sub> solar cells
fabricated from thermal evaporation. The rationale to choose thermal
evaporation for Sb<sub>2</sub>Se<sub>3</sub> film deposition was first
discussed, followed by detailed characterization of Sb<sub>2</sub>Se<sub>3</sub> film deposited onto FTO with different substrate temperatures.
We then studied the optical absorption, photosensitivity, and band
position of Sb<sub>2</sub>Se<sub>3</sub> film, and finally a prototype
photovoltaic device FTO/Sb<sub>2</sub>Se<sub>3</sub>/CdS/ZnO/ZnO:Al/Au
was constructed, achieving an encouraging 2.1% solar conversion efficiency
The glycosyl hydrolase activity of <i>S. pneumoniae</i> GHIP.
<p>(A) <i>S. pneumoniae</i> GHIP (green) superimposed on its homolog from <i>Bacillus anthracis</i>, PlyB (cyan) (PDB code 2NW0). The magenta arrow points toward the major difference, where the N-terminal regions form a helix (α1) which is absent in PlyB. The sticks represent the four key acidic residues to their enzyme activities, inculding Asp56, Asp154, Glu156, and Asp245 of GHIP and Asp6, Asp90, Asp92, and Asp171 of PlyB. (B) The enlarged image of the four key residues at the active site. Close up view of <i>S. pneumoniae</i> GHIP showing overlay of key acidic residues: Asp56, Asp154, Glu156, and Asp245 of GHIP and Asp6, Asp90, Asp92, and Asp171 of PlyB, which exhibit similar locations and orientations. (C) The month of TIM barrel in <i>S. pneumoniae</i> GHIP is active site which contains 14 residues in sticks, including Asp56, Ser58, Ser84, Tyr121, Tyr123, Glu154, Glu156, Asp157, Tyr185, Tyr209, Asp212, Ser233, Asp243 and Asp245. The GHIP structure is shown in the context of a transparent (gray) surface. (D) The enlarged image of the 14 residues at the active site and the background is the electrostatic potential surface of <i>S. pneumoniae</i> GHIP. Saturated red indicates Φ<−10 kiloteslas/e, and saturated blue indicates Φ>10 kiloteslas/e, T = 20°C. (E) & (F) The temperature and pH activity analyses of <i>S. pneumoniae</i> GHIP. Peptidoglycan hydrolase activity was measured at various pH (4.0 to 8.0) and temperatures (25 to 45°C) ranges as described in the Materials and Methods. (G) Hydrolase activity of <i>S. pneumoniae</i> GHIP on PNP-(GlcNAc)<sub>5</sub>. Lane 1 represents the positive control using HEWL (egg white lysozyme); lane 2 is native GHIP; lanes 3–6 are active-site mutants D56A, D154A, E156A, and D245A, respectively. Asterisks denote values significantly different from the wild-type strain by Student’s <i>t</i>-test (*, P<0.05; **, P<0.01).</p
Animal and colonization experiments testing <i>S. pneumoniae</i> GHIP function.
<p>(A) & (B) The effect of the <i>GHIP</i> deletion mutation on virulence. Groups of 18 BALB/c mice were intranasally or intraperitoneally challenged with 1.0 × 10<sup>8</sup> and 1.0 × 10<sup>3</sup> CFU, respectively, of D39 or the isogenic <i>ΔGHIP</i> mutant. Each datum point represents one mouse. Solid line, wild-type D39; dotted line, <i>ΔGHIP</i> mutant. Asterisks denote values significantly different from wild-type by Student’s <i>t-</i>test (*, P<0.05). (C) The effect of the <i>GHIP</i> deletion mutation on bacteria recovered from nasopharynxes, lungs, and blood samples of BALB/c mice after intranasal challenge. BALB/c mice were intranasally challenged with either wild-type D39 or the <i>ΔGHIP</i> mutant at 1.0 × 10<sup>8 </sup>CFU/mouse. At 12, 24, and 36 h post-infection, six mice from each group were scarified, and the number of recovered bacteria was determined by plating on blood agar. Results are expressed as log<sub>10</sub> CFUs per gram tissue and represent individually recorded values for each mouse. Horizontal bars represent geometric means. Asterisks denote values significantly different from wild-type by Student’s <i>t-</i>test (*, P<0.05).</p
Amino-acid sequence alignment comparing homologs of <i>S.pneumoniae</i> GHIP.
<p>Sequence alignment of five streptococcus SpGHIP homologs, including SpGHIP from <i>Streptococcus pneumoniae</i> TIGR4 (accession number NP_345466), <i>Streptococcus gordonii</i> (accession number YP_001450056), <i>Streptococcus sanguinis</i> (accession number EGD37983), <i>Streptococcus pyogenes</i> (accession number NP_268858), <i>Streptococcus suis</i> (accession number EHC03477), and PlyB from the BcpI phage (PDB code 2NW0). Identical and similar residues are shown in <i>cyan</i> and <i>yellow</i>, respectively. Secondary structure elements of SpGHIP are shown above the sequences. Residues are numbered according to the SpGHIP sequence. Sequence alignment was generated using ClustalW.</p
Adherence and invasion of the <i>ΔGHIP</i> mutant into host cells <i>in vitro</i>.
<p>(A) & (B) <i>S. pneumoniae</i> type R6 and their isogenic R6Δ<i>GHIP</i> mutants were examined for adherence to A549 and CNE2 cells, respectively. (C) & (D) <i>S. pneumoniae</i> type R6 and their isogenic R6Δ<i>GHIP</i> were examined for invasion into A549 and CNE2 cells, respectively. Infection experiments were conducted for 4 h at 37°C. Wild-type <i>S. pneumoniae</i> and GHIP-deficient mutants were generated to be devoid of pneumolysin in order to avoid destruction of the monolayer in the <i>in vitro</i> infection system. Scoring the number of adherent and invasive bacteria indicate that adhesion and invasion are substantially reduced for GHIP-deficient <i>S. pneumoniae</i>. Results are presented as the mean ± standard deviation for at least three independent experiments. Asterisks and triangles denote values significantly different from wild-type by Student’s <i>t</i>-test (**, P<0.01; ▴, P<0.05).</p
X-ray data collection and refinement statistics of <i>S. pneumoniae</i> GHIP.
a<p>Values in parentheses are for the highest-resolution shells.</p>b<p>R<sub>merge</sub> = ∑(I−<i>)/∑I, where I is the observed intensity and <i> is the statistically weighted average intensity of multiple symmetry related observation.</i></i></p><i><i>c<p>R<sub>factor</sub>: R = ∑||F<sub>calc</sub>|−|F<sub>obs</sub>|/∑|F<sub>obs</sub>|, where F<sub>calc</sub> and F<sub>obs</sub> are the calculated and observed structure factors, respectively. R<sub>free</sub> is calculated the same as R<sub>factor</sub> but from a test set containing 5% of data excluded from the refinement calculation.</p>d<p>rmsd, root-mean-square deviation from ideal geometry.</p></i></i