Structural basis for inhibition of complement C5 by the SSL7 protein from Staphylococcus aureus

AbstractTransporters play vital roles in acquiring antimicrobial resistance among pathogenic bacteria. In this study, we report the X-ray structure of NorC, a 14-transmembrane major facilitator superfamily member that is implicated in fluoroquinolone resistance in drug-resistant Staphylococcus aureus strains, at a resolution of 3.6 Å. The NorC structure was determined in complex with a single-domain camelid antibody that interacts at the extracellular face of the transporter and stabilizes it in an outward-open conformation. The complementarity determining regions of the antibody enter and block solvent access to the interior of the vestibule, thereby inhibiting alternating-access. NorC specifically interacts with an organic cation, tetraphenylphosphonium, although it does not demonstrate an ability to transport it. The interaction is compromised in the presence of NorC-antibody complex, consequently establishing a strategy to detect and block NorC and related transporters through the use of single-domain camelid antibodies.

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Structural Basis for Linezolid Binding Site Rearrangement in the Staphylococcus aureus Ribosome

mBio ◽

10.1128/mbio.00395-17 ◽

2017 ◽

Vol 8 (3) ◽

Cited By ~ 14

Author(s):

Matthew J. Belousoff ◽

Zohar Eyal ◽

Mazdak Radjainia ◽

Tofayel Ahmed ◽

Rebecca S. Bamert ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Structural Information ◽

Structural Basis ◽

Common Mechanism ◽

Resistance Mutations ◽

Antimicrobial Drugs ◽

Linezolid Resistance ◽

Drug Modification ◽

Antibiotic Resistance Mutations ◽

Shed Light

ABSTRACT An unorthodox, surprising mechanism of resistance to the antibiotic linezolid was revealed by cryo-electron microscopy (cryo-EM) in the 70S ribosomes from a clinical isolate of Staphylococcus aureus. This high-resolution structural information demonstrated that a single amino acid deletion in ribosomal protein uL3 confers linezolid resistance despite being located 24 Å away from the linezolid binding pocket in the peptidyl-transferase center. The mutation induces a cascade of allosteric structural rearrangements of the rRNA that ultimately results in the alteration of the antibiotic binding site. IMPORTANCE The growing burden on human health caused by various antibiotic resistance mutations now includes prevalent Staphylococcus aureus resistance to last-line antimicrobial drugs such as linezolid and daptomycin. Structure-informed drug modification represents a frontier with respect to designing advanced clinical therapies, but success in this strategy requires rapid, facile means to shed light on the structural basis for drug resistance (D. Brown, Nat Rev Drug Discov 14:821–832, 2015, https://doi.org/10.1038/nrd4675 ). Here, detailed structural information demonstrates that a common mechanism is at play in linezolid resistance and provides a step toward the redesign of oxazolidinone antibiotics, a strategy that could thwart known mechanisms of linezolid resistance. IMPORTANCE The growing burden on human health caused by various antibiotic resistance mutations now includes prevalent Staphylococcus aureus resistance to last-line antimicrobial drugs such as linezolid and daptomycin. Structure-informed drug modification represents a frontier with respect to designing advanced clinical therapies, but success in this strategy requires rapid, facile means to shed light on the structural basis for drug resistance (D. Brown, Nat Rev Drug Discov 14:821–832, 2015, https://doi.org/10.1038/nrd4675 ). Here, detailed structural information demonstrates that a common mechanism is at play in linezolid resistance and provides a step toward the redesign of oxazolidinone antibiotics, a strategy that could thwart known mechanisms of linezolid resistance.

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Convergent Evolution of Neutralizing Antibodies to Staphylococcus aureus γ-Hemolysin C That Recognize an Immunodominant Primary Sequence-Dependent B-Cell Epitope

mBio ◽

10.1128/mbio.00460-20 ◽

2020 ◽

Vol 11 (3) ◽

Cited By ~ 2

Author(s):

David N. Hernandez ◽

Kayan Tam ◽

Bo Shopsin ◽

Emily E. Radke ◽

Karen Law ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Phage Display ◽

B Cell ◽

Neutralizing Antibodies ◽

Ex Vivo ◽

Cell Epitope ◽

Phage Display Library ◽

Structural Basis ◽

Peptide Fragments ◽

Content Type

ABSTRACT Staphylococcus aureus infection is a major public health threat in part due to the spread of antibiotic resistance and repeated failures to develop a protective vaccine. Infection is associated with production of virulence factors that include exotoxins that attack host barriers and cellular defenses, such as the leukocidin (Luk) family of bicomponent pore-forming toxins. To investigate the structural basis of antibody-mediated functional inactivation of Luk toxins, we generated a panel of murine monoclonal antibodies (MAbs) that neutralize host cell killing by the γ-hemolysin HlgCB. By biopanning these MAbs against a phage-display library of random Luk peptide fragments, we identified a small subregion within the rim domain of HlgC as the epitope for all the MAbs. Within the native holotoxin, this subregion folds into a conserved β-hairpin structure, with exposed key residues, His252 and Tyr253, required for antibody binding. On the basis of the phage-display results and molecular modeling, a 15-amino-acid synthetic peptide representing the minimal epitope on HlgC (HlgC241-255) was designed, and preincubation with this peptide blocked antibody-mediated HIgCB neutralization. Immunization of mice with HlgC241-255 or the homologous LukS246-260 subregion peptide elicited serum antibodies that specifically recognized the native holotoxin subunits. Furthermore, serum IgG from patients who were convalescent for invasive S. aureus infection showed neutralization of HlgCB toxin activity ex vivo, which recognized the immunodominant HlgC241-255 peptide and was dependent on His252 and Tyr253 residues. We have thus validated an efficient, rapid, and scalable experimental workflow for identification of immunodominant and immunogenic leukotoxin-neutralizing B-cell epitopes that can be exploited for new S. aureus-protective vaccines and immunotherapies.

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Author Correction: Structural basis of cell wall peptidoglycan amidation by the GatD/MurT complex of Staphylococcus aureus

Scientific Reports ◽

10.1038/s41598-018-37082-9 ◽

2018 ◽

Vol 8 (1) ◽

Author(s):

Erik R. Nöldeke ◽

Lena M. Muckenfuss ◽

Volker Niemann ◽

Anna Müller ◽

Elena Störk ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Cell Wall ◽

Structural Basis ◽

Cell Wall Peptidoglycan

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Structural basis for the O-acetyltransferase function of the extracytoplasmic domain of OatA from Staphylococcus aureus

Journal of Biological Chemistry ◽

10.1074/jbc.ra120.013108 ◽

2020 ◽

Vol 295 (24) ◽

pp. 8204-8213 ◽

Cited By ~ 1

Author(s):

Carys S. Jones ◽

David Sychantha ◽

P. Lynne Howell ◽

Anthony J. Clarke

Keyword(s):

Staphylococcus Aureus ◽

Biochemical Characterization ◽

Multidrug Resistant ◽

Catalytic Triad ◽

Structural Basis ◽

Gram Positive ◽

Gram Positive Bacteria ◽

Hydrolase Fold ◽

Acetyl Transfer ◽

Human Infections

Many bacteria possess enzymes that modify the essential cell-wall polymer peptidoglycan by O-acetylation. This modification occurs in numerous Gram-positive pathogens, including methicillin-resistant Staphylococcus aureus, a common cause of human infections. O-Acetylation of peptidoglycan protects bacteria from the lytic activity of lysozyme, a mammalian innate immune enzyme, and as such is important for bacterial virulence. The O-acetylating enzyme in Gram-positive bacteria, O-acetyltransferase A (OatA), is a two-domain protein consisting of an N-terminal integral membrane domain and a C-terminal extracytoplasmic domain. Here, we present the X-ray crystal structure at 1.71 Å resolution and the biochemical characterization of the C-terminal domain of S. aureus OatA. The structure revealed that this OatA domain adopts an SGNH-hydrolase fold and possesses a canonical catalytic triad. Site-specific replacement of active-site amino acids revealed the presence of a water-coordinating aspartate residue that limits esterase activity. This residue, although conserved in staphyloccocal OatA and most other homologs, is not present in the previously characterized streptococcal OatA. These results provide insights into the mechanism of acetyl transfer in the SGNH/GDSL hydrolase family and highlight important evolutionary differences between homologous OatA enzymes. Furthermore, this study enhances our understanding of PG O-acetyltransferases, which could guide the development of novel antibacterial drugs to combat infections with multidrug-resistant bacterial pathogens.

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Crystal structure of Staphylococcus aureus exfoliative toxin D-like protein: Structural basis for the high specificity of exfoliative toxins

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2015.08.083 ◽

2015 ◽

Vol 467 (1) ◽

pp. 171-177 ◽

Cited By ~ 3

Author(s):

Ricardo B. Mariutti ◽

Tatiana A.C.B. Souza ◽

Anwar Ullah ◽

Icaro P. Caruso ◽

Fábio R. de Moraes ◽

...

Keyword(s):

Crystal Structure ◽

Staphylococcus Aureus ◽

High Specificity ◽

Structural Basis ◽

Exfoliative Toxin

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Structural basis for evasion of IgA immunity by Staphylococcus aureus revealed in the complex of SSL7 with Fc of human IgA1

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.0706028104 ◽

2007 ◽

Vol 104 (38) ◽

pp. 15051-15056 ◽

Cited By ~ 78

Author(s):

P. A. Ramsland ◽

N. Willoughby ◽

H. M. Trist ◽

W. Farrugia ◽

P. M. Hogarth ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Structural Basis

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Crystal structure of CTP:glycerol-3-phosphate cytidylyltransferase from Staphylococcus aureus: Examination of structural basis for kinetic mechanism

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics ◽

10.1016/j.bbapap.2005.10.015 ◽

2006 ◽

Vol 1764 (1) ◽

pp. 63-69 ◽

Cited By ~ 12

Author(s):

Desiree H. Fong ◽

Veronica C.-N. Yim ◽

Michael A. D'Elia ◽

Eric D. Brown ◽

Albert M. Berghuis

Keyword(s):

Crystal Structure ◽

Staphylococcus Aureus ◽

Kinetic Mechanism ◽

Structural Basis

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A Single, Engineered Protein Therapeutic Agent Neutralizes Exotoxins from Both Staphylococcus aureus and Streptococcus pyogenes

Clinical and Vaccine Immunology ◽

10.1128/cvi.00277-10 ◽

2010 ◽

Vol 17 (11) ◽

pp. 1781-1789 ◽

Cited By ~ 16

Author(s):

Ningyan Wang ◽

Daiva M. Mattis ◽

Eric J. Sundberg ◽

Patrick M. Schlievert ◽

David M. Kranz

Keyword(s):

Staphylococcus Aureus ◽

T Cell ◽

Streptococcus Pyogenes ◽

Sequence Similarity ◽

Large Fraction ◽

Cross Reactivity ◽

Structural Basis ◽

Cell Repertoire ◽

High Affinity ◽

The Cross

ABSTRACT Staphylococcus aureus and Streptococcus pyogenes secrete exotoxins that act as superantigens, proteins that cause hyperimmune reactions by binding the variable domain of the T-cell receptor beta chain (Vβ), leading to stimulation of a large fraction of the T-cell repertoire. To develop potential neutralizing agents, we engineered Vβ mutants with high affinity for the superantigens staphylococcal enterotoxin B (SEB), SEC3, and streptococcal pyrogenic exotoxin A (SpeA). Unexpectedly, the high-affinity Vβ mutants generated against SEB cross-reacted with SpeA to a greater extent than they did with SEC3, despite greater sequence similarity between SEB and SEC3. Likewise, the Vβ mutants generated against SpeA cross-reacted with SEB to a greater extent than with SEC3. The structural basis of the high affinity and cross-reactivity was examined by single-site mutational analyses. The cross-reactivity seems to involve only one or two toxin residues. Soluble forms of the cross-reactive Vβ regions neutralized both SEB and SpeA in vivo, suggesting structure-based strategies for generating high-affinity neutralizing agents that can cross-react with multiple exotoxins.

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