Electron microscopy and computational studies of Ebh, a giant cell-wall-associated protein from Staphylococcus aureus

Sakamoto, Sou; Tanaka, Yoshikazu; Tanaka, Isao; Takei, Toshiaki; Yu, Jian; Kuroda, Makoto; Yao, Min; Ohta, Toshiko; Tsumoto, Kouhei

doi:10.1016/j.bbrc.2008.08.117

Cited by 8 publications

(7 citation statements)

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“…The ebh gene is the largest open reading frame on the S. aureus chromosome at 33 kb, effectively comprising 1% of the genome [19]. Ebh, also called Giant Staphylococcal Surface Protein (GSSP), is predicted to be membrane anchored and protruding from the cell surface in a fiber-like manner [36], [37]. The protein contains numerous sugar binding and albumin binding repeat domains that are thought to be important for binding extracellular matrix proteins [19].…”

Section: Resultsmentioning

confidence: 99%

The Staphylococcus aureus ArlRS Two-Component System Is a Novel Regulator of Agglutination and Pathogenesis

et al. 2013

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Staphylococcus aureus is a prominent bacterial pathogen that is known to agglutinate in the presence of human plasma to form stable clumps. There is increasing evidence that agglutination aids S. aureus pathogenesis, but the mechanisms of this process remain to be fully elucidated. To better define this process, we developed both tube based and flow cytometry methods to monitor clumping in the presence of extracellular matrix proteins. We discovered that the ArlRS two-component system regulates the agglutination mechanism during exposure to human plasma or fibrinogen. Using divergent S. aureus strains, we demonstrated that arlRS mutants are unable to agglutinate, and this phenotype can be complemented. We found that the ebh gene, encoding the Giant Staphylococcal Surface Protein (GSSP), was up-regulated in an arlRS mutant. By introducing an ebh complete deletion into an arlRS mutant, agglutination was restored. To assess whether GSSP is the primary effector, a constitutive promoter was inserted upstream of the ebh gene on the chromosome in a wildtype strain, which prevented clump formation and demonstrated that GSSP has a negative impact on the agglutination mechanism. Due to the parallels of agglutination with infective endocarditis development, we assessed the phenotype of an arlRS mutant in a rabbit combined model of sepsis and endocarditis. In this model the arlRS mutant displayed a large defect in vegetation formation and pathogenesis, and this phenotype was partially restored by removing GSSP. Altogether, we have discovered that the ArlRS system controls a novel mechanism through which S. aureus regulates agglutination and pathogenesis.

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Section: Resultsmentioning

confidence: 99%

The Staphylococcus aureus ArlRS Two-Component System Is a Novel Regulator of Agglutination and Pathogenesis

et al. 2013

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“…X-ray crystallography revealed FIVAR-GA as double (FIVAR)-and triple (GA)-helical bundle structures with an elongated shape (24,29). Other work reported that S. aureus 8325-4 ebh mutants display increased sensitivity to teicoplanin (a glycopeptide antibiotic), osmotic stress, and Triton X-100 (23).…”

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confidence: 99%

The Giant Protein Ebh Is a Determinant of Staphylococcus aureus Cell Size and Complement Resistance

Cheng

Missiakas

Schneewind

2013

Journal of Bacteriology

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Staphylococcus aureus USA300, the clonal type associated with epidemic community-acquired methicillin-resistant S. aureus (MRSA) infections, displays the giant protein Ebh on its surface. Mutations that disrupt the ebh reading frame increase the volume of staphylococcal cells and alter the cross wall, a membrane-enclosed peptidoglycan synthesis and assembly compartment. S. aureus ebh variants display increased sensitivity to oxacillin (methicillin) as well as susceptibility to complement-mediated killing. Mutations in ebh are associated with reduced survival of mutant staphylococci in blood and diminished virulence in mice. We propose that Ebh, following its secretion into the cross wall, contributes to the characteristic cell growth and envelope assembly pathways of S. aureus, thereby enabling complement resistance and the pathogenesis of staphylococcal infections.

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“…Additionally, a previous report that purified 10 of the large sugar and albumin binding domains from Escherichia coli suggests the protein protrudes from the cell surface in a fiber like manner [67]. The N-terminal signal sequence and C-terminal anchor domain encoded in the ebh sequence also provide strong predictive evidence that the protein is surface exposed.…”

Section: Future Directions To Determine Ebh Productionmentioning

confidence: 86%

“…The ebh gene is the largest open reading frame on the S. aureus chromosome at 33 kb, effectively comprising 1% of the genome [66]. The ebh gene encodes a surface bound protein, predicted to protrude from the membrane in a fiber-like manner [67,68]. Preliminary studies predict the protein binds host matrix components via the large number of sugar and albumin binding repeat domains maintained within the sequence [66].…”

Section: Discussionmentioning

confidence: 99%

“…The shear size of the Ebh protein, at 1.1Mda, could act to physically separate two bacterial cells, keeping the MSCRAMMs far enough apart that they are unable to come into contact with the same ECM protein. Previous studies suggest Ebh is a surface anchored protein that protrudes from the membrane in a fiberlike manner [66][67][68][69]. This rod-like structure is thought to have some flexibility [68].…”

Section: Future Directions To Determine the Mechanism By Which Ebh Inmentioning

confidence: 99%

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The two-component system, ArlRS, regulates agglutination and pathogenesis in Staphylococcus aureus

Walker¹

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____________________________________________________ Bradley Jones ii To my parents, for encouraging me to work towards my goals and providing with the support I needed to reach them iii ACKNOWLEDGEMENTS I would like to extend my deepest thanks to the family, friends, labmates, teachers, and professors who have played an instrumental role in helping me reach my goals. You have instilled in me a love of science and learning and provided the support and training necessary to be a successful research-based scientist. Thank you to my thesis committee members for providing helpful insights and advice on my project. My project would not have been as successful without your guidance. To my labmates in the Horswill lab, you are all incredibly talented investigators and even better friends. The lab bonding time, on trips, at conferences, and during coffee breaks, spent with all of you made my graduate school experience that much better. To Alex, thank you for providing me the freedom on my project to pursue the questions and experiments that were of interest to me and the opportunity to present my work at so many great conferences. To Jeff and Cherie, thank you for all your hard work trying to keep the graduate students in line and the lab running smoothly. To Jeff Boyd, your help making mutations has beenan incredible benefit to my project and allowed us to aim for an outstanding publication.To Laynie, I appreciate your help with qPCR and providing all of the equipment and software to enable me to do the qPCR experiments. Your insights into how bees make honey and the differences in hot peppers have made me a more well rounded scientist.To Bill Nauseef and Jamie Schwartz, thank you for trying to teach me the intricacies of the immune system and how to use a flow cytometer. To Pat Schlievert, Adam, and Wilmara, thank you for all your help with the animal models. I greatly appreciate all the hard work you put into helping me with my project. To Jovanka Voyich and the Voyich iv lab, thank you for being fabulous. Having a colleague to collaborate with on a difficult project made things much easier and kept my spirits up when things were challenging. Thank you for taking the time and having the patience to teach me how to do mouse experiments. To Blake, thank you for trying to teach me statistics and bioinformatics. I will never understand them quite the way you do, but as long as you are there to check my work I don't really need to. Lastly, I would like to thank the Hultgren Lab, specifically Jerry Pinkner and Scott Hultgren, for giving me the opportunity, as a senior in high school, to do research in your lab. Your encouragement and confidence in my ability to do hypothesis-based research instilled in me a love of science and a conviction to be successful in all of my endeavors. The support of many people over many years allowed me to be successful and achieve my goals. v ABSTRACT Staphylococcus aureus is defined by its ability to agglutinate during exposure to human blood plasma. Although agglutination has long correlated...

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Electron microscopy and computational studies of Ebh, a giant cell-wall-associated protein from Staphylococcus aureus

Cited by 8 publications

References 13 publications

The Staphylococcus aureus ArlRS Two-Component System Is a Novel Regulator of Agglutination and Pathogenesis

The Staphylococcus aureus ArlRS Two-Component System Is a Novel Regulator of Agglutination and Pathogenesis

The Giant Protein Ebh Is a Determinant of Staphylococcus aureus Cell Size and Complement Resistance

The two-component system, ArlRS, regulates agglutination and pathogenesis in Staphylococcus aureus

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