Structural and Functional Characterization of the Streptococcus pneumoniae RrgB Pilus Backbone D1 Domain

Gentile, Michela; Melchiorre, Sara; Emolo, Carla; Moschioni, Monica; Gianfaldoni, Claudia; Pancotto, Laura; Ferlenghi, Ilaria; Scarselli, María; Pansegrau, Werner; Veggi, Daniele; Merola, Marcello; Cantini, Francesca; Ruggiero, Paolo; Banci, Lucia; Masignani, Vega

doi:10.1074/jbc.m110.202739

Cited by 22 publications

(30 citation statements)

References 55 publications

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“…Like some of the three‐domain backbone pilins (FimA, GBS80), a loss of the D1 domain is also noticed in the four‐domain pilins . However, a solution structure of D1 domain of RrgB with no intramolecular isopeptide bond has been reported . A complete structure containing all four domains, and including a sorting motif, has also been reported for RrgB .…”

Section: Backbone Pilinsmentioning

confidence: 96%

“…Several pilin structures contain metal ions, which might play a structural role. The available crystal structures for backbone pilins are Spy0128 and T6 from S. pyogenes , SpaA and SpaD from C. diphtheriae , BcpA from B. cereus , FimA and FimP from A. oris , GBS80, BP‐2a, and BP‐2b from S. agalactiae , and RrgB from S. pneumonia (Fig. ).…”

Section: Backbone Pilinsmentioning

confidence: 99%

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Pilins in gram‐positive bacteria: A structural perspective

Krishnan

2015

IUBMB Life

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Pilins or fimbrilins are a class of proteins found in bacterial surface pilus, a hair-like surface appendage. Both the Gramnegative and -positive bacteria produce pilins to assemble pili on their cell-surface for different purposes including adherence, twitching motility, conjugation, immunomodulation, biofilm formation, and electron transfer. Immunogenic properties of the pilins make them attractive vaccine candidates. The polymerized pilins play a key role in the initiation of host adhesion, which is a critical step for bacterial colonization and infection. Because of their key role in adhesion and exposure on the cell surface, targeting the pilins-mediated adhesion (anti-adhesion therapy) is also seen as a promising alternative approach for preventing and treating bacterial infections, one that may overcome their ever-increasing repertoires of resistance mechanisms. Individual pilins interact with each other non-covalently to assemble the pilus fiber with the help of associated proteins like chaperones and Usher in Gramnegative bacteria. In contrast, the pilins in Gram-positive bacteria often connect with each other covalently, with the help of sortases. Certain unique structural features present on the pilins distinguish them from one another across different bacterial strains, and these dictate their cellular targets and functions. While the structure of pilins has been extensively studied in Gram-negative pathogenic bacteria, the pilins in Gram-positive pathogenic bacteria have been in only during the last decade. Recently, the discovery of pilins in nonpathogenic bacteria, such as Lactobacillus rhamnosus GG, has received great attention, though traditionally the attention was on pathogenic bacteria. This review summarizes and discusses the current structural knowledge of pilins in Gram-positive bacteria with emphasis on those pilins which are sortase substrates. V C 2015 IUBMB Life, 67(7): [533][534][535][536][537][538][539][540][541][542][543] 2015

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Section: Backbone Pilinsmentioning

confidence: 96%

Section: Backbone Pilinsmentioning

confidence: 99%

Pilins in gram‐positive bacteria: A structural perspective

Krishnan

2015

IUBMB Life

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“…Different structural models of the type 1 pilus have been proposed (12,16,17,34), but in general, it is agreed that the RrgB protein forms the backbone, to which are attached two ancillary proteins, RrgA and RrgC, which are thought to be an adhesin and anchor to the cell wall, respectively. Prior to universal immunization with pneumococcal conjugate vaccine Prevnar in the United States, the type 1 pilus was found predominantly in serotypes that were included in the vaccine (5).…”

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confidence: 99%

Expression of the Type 1 Pneumococcal Pilus Is Bistable and Negatively Regulated by the Structural Component RrgA

et al. 2011

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The pneumococcal type 1 pilus, which is present in 25 to 30% of clinical isolates, has been associated with increased adherence and inflammatory responses and is being evaluated as a potential vaccine candidate. Here we show that expression of the pilus is bistable as a result of the molecular interaction between the transcription activator RrlA and a structural component of the pilus called RrgA. Sampling various clinical pneumococcal isolates that harbor the type 1 pilus-encoding islet, we show that distinct populations of cells can be identified with either undetectable or prominent pilus expression. When these two populations are separated and regrown in liquid medium, they are phenotypically different: the nonexpressing population reverts to the previous bimodal distribution, whereas the expressing population retains the same high level of pilus expression. Controlled exogenous expression of the regulatory pilus gene rlrA in a strain from which the endogenous version has been deleted increases pilus expression steadily, suggesting that the bistable expression of the pilus observed in wild-type cells is dependent on the native rlrA promoter. Finally, we demonstrate that RrgA is a negative regulator of pilus expression and that this repression is likely mediated through direct interaction with RlrA. We conclude that type 1 pilus expression in pneumococcus exhibits a bistable phenotype, which is dependent upon the molecular interplay between the RlrA and RrgA proteins. We suggest that this flexibility in expression may assist adaptation to a range of immune conditions, such as evasion of antipilus antibodies, within potential hosts.

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“…The shuttle plasmid pMU1328_Pc_wchA was obtained as described below. Briefly, the wchA gene of the AP425 strain and the constitutive promoter of erythromycin (Pc) (Gentile et al, 2011) were amplified with the primers wchA_for (59-GTGCGTGGATCCATGGATGAAAAAGGA-TTGAAAATT-39) and wchA_rev (59-CAGCGTGGATCCTCACTT-CGCCCCTTCTCTCATAAA-39), and Pc_for (59-GTGCGTGAATT-CGAAACAGCAAAGAATGGCGGAAAC-39) and Pc_rev (59-CAGC-GTGGATCCGTAATCACTCCTTCTTAATTACAA-39), respectively. The two PCR products, digested with BamHI and EcoRI/BamHI restriction enzymes, respectively, were cloned into pMU1328, containing an erythromycin-resistance marker (Achen et al, 1986).…”

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confidence: 99%

Point mutations in wchA are responsible for the non-typability of two invasive Streptococcus pneumoniae isolates

et al. 2012

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Non-typable Streptococcus pneumoniae (NTPn) strains are typically isolated from nasopharyngeal carriage or from conjunctivitis. Since the isolation of NTPn from invasive disease is rare, we characterized the genetic basis of the non-typability of two isolates obtained in Italy from two cases of bacteraemic pneumonia. MLST revealed that both NTPn belonged to ST191, which, according to the MLST database, is associated with serotype 7F. Sequencing of the capsular locus (cps) confirmed the presence of a 7F cps in both strains and revealed the existence of distinct single point mutations in the wchA gene (a glycosyltransferase), both leading to the translation of proteins truncated at the C terminus. To verify that these mutations were responsible for the non-typability of the isolates, a functional 7F WchA was overexpressed in both NTPn. The two NTPn along with their WchA-overexpressing derivatives were analysed by transmission electron microscopy and by high-resolution magic angle spinning NMR spectroscopy. Both NTPn were devoid of a polysaccharide capsule, and WchA overexpression was sufficient to restore the assembly of a serotype 7F capsule on the surface of the two NTPn. In conclusion, we identified two new naturally occurring point mutations that lead to non-typability in the pneumococcus, and demonstrated that WchA is essential for the biosynthesis of the serotype 7F capsule. INTRODUCTIONThe Gram-positive pathogen Streptococcus pneumoniae is a major cause of community-acquired pneumonia, as well as upper respiratory tract infections such as acute otitis media and sinusitis, and invasive diseases such as meningitis, bacteraemia and endocarditis. However, S. pneumoniae is also a commensal of the upper respiratory tract, especially of young children, who represent the reservoir for pneumococcal transmission within the community.The ability of the pneumococcus to invade sterile sites is related to the expression of a polysaccharide capsule, which provides a barrier against host cell-mediated phagocytosis and allows bacterial persistence in the blood (Brown et al., 1983). In addition, capsular polysaccharides (CPSs) are immunogenic and antibodies against CPSs provide protection against pneumococcal disease; therefore, all of the current pneumococcal vaccines are based on a combination of different CPSs, either unconjugated or conjugated to a carrier protein (Gladstone et al., 2011).To date, 93 capsular types (serotypes) have been described on the basis of their different genetic, biochemical and antigenic properties (Bentley et al., 2006;Calix & Nahm, 2010;Park et al., 2007). Each pneumococcal serotype corresponds to a distinct CPS, synthesized on the bacterial surface by enzymes which are encoded within a single capsule locus (cps), with a few notable exceptions (Llull et al., 1999). The sequencing of different cps loci has revealed the presence of a core set of genes common to all capsular types, namely wzg, wzh, wzd and wze (also known as cpsA, B, C and D), and of additional genes that are CPS-specific (Bentley...

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Structural and Functional Characterization of the Streptococcus pneumoniae RrgB Pilus Backbone D1 Domain

Cited by 22 publications

References 55 publications

Pilins in gram‐positive bacteria: A structural perspective

Pilins in gram‐positive bacteria: A structural perspective

Expression of the Type 1 Pneumococcal Pilus Is Bistable and Negatively Regulated by the Structural Component RrgA

Point mutations in wchA are responsible for the non-typability of two invasive Streptococcus pneumoniae isolates

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