Journal articles: 'Autotransporters'

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Jain, Sumita, Peter van Ulsen, Inga Benz, M. Alexander Schmidt, Rachel Fernandez, Jan Tommassen, and Marcia B. Goldberg. "Polar Localization of the Autotransporter Family of Large Bacterial Virulence Proteins." Journal of Bacteriology 188, no. 13 (July 1, 2006): 4841–50. http://dx.doi.org/10.1128/jb.00326-06.

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ABSTRACT Autotransporters are an extensive family of large secreted virulence-associated proteins of gram-negative bacteria. Secretion of such large proteins poses unique challenges to bacteria. We demonstrate that autotransporters from a wide variety of rod-shaped pathogens, including IcsA and SepA of Shigella flexneri, AIDA-I of diffusely adherent Escherichia coli, and BrkA of Bordetella pertussis, are localized to the bacterial pole. The restriction of autotransporters to the pole is dependent on the presence of a complete lipopolysaccharide (LPS), consistent with known effects of LPS composition on membrane fluidity. Newly synthesized and secreted BrkA is polar even in the presence of truncated LPS, and all autotransporters examined are polar in the cytoplasm prior to secretion. Together, these findings are consistent with autotransporter secretion occurring at the poles of rod-shaped gram-negative organisms. Moreover, NalP, an autotransporter of spherically shaped Neisseria meningitidis contains the molecular information to localize to the pole of Escherichia coli. In N. meningitidis, NalP is secreted at distinct sites around the cell. These data are consistent with a model in which the secretion of large autotransporters occurs via specific conserved pathways located at the poles of rod-shaped bacteria, with profound implications for the underlying physiology of the bacterial cell and the nature of bacterial pathogen-host interactions.

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Jain, Sumita, and Marcia B. Goldberg. "Requirement for YaeT in the Outer Membrane Assembly of Autotransporter Proteins." Journal of Bacteriology 189, no. 14 (May 18, 2007): 5393–98. http://dx.doi.org/10.1128/jb.00228-07.

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ABSTRACT Autotransporters constitute the largest group of secreted proteins in gram-negative bacteria. Autotransporter secretion involves the insertion of a carboxy-terminal beta barrel into and the translocation of an amino-terminal domain across the outer membrane. Here, we demonstrate that secretion of autotransporters from several organisms requires the outer membrane assembly factor YaeT.

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Lane, M. Chelsea, Jonathan D. Lenz, and Virginia L. Miller. "Proteolytic processing of the Yersinia pestis YapG autotransporter by the omptin protease Pla and the contribution of YapG to murine plague pathogenesis." Journal of Medical Microbiology 62, no. 8 (August 1, 2013): 1124–34. http://dx.doi.org/10.1099/jmm.0.056275-0.

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Autotransporter protein secretion represents one of the simplest forms of secretion across Gram-negative bacterial membranes. Once secreted, autotransporter proteins either remain tethered to the bacterial surface or are released following proteolytic cleavage. Autotransporters possess a diverse array of virulence-associated functions such as motility, cytotoxicity, adherence and autoaggregation. To better understand the role of autotransporters in disease, our research focused on the autotransporters of Yersinia pestis, the aetiological agent of plague. Y. pestis strain CO92 has nine functional conventional autotransporters, referred to as Yaps for Yersinia autotransporter proteins. Three Yaps have been directly implicated in virulence using established mouse models of plague infection (YapE, YapJ and YapK). Whilst previous studies from our laboratory have shown that most of the CO92 Yaps are cell associated, YapE and YapG are processed and released by the omptin protease Pla. In this study, we identified the Pla cleavage sites in YapG that result in many released forms of YapG in Y. pestis, but not in the evolutionarily related gastrointestinal pathogen, Yersinia pseudotuberculosis, which lacks Pla. Furthermore, we showed that YapG does not contribute to Y. pestis virulence in established mouse models of bubonic and pneumonic infection. As Y. pestis has a complex life cycle involving a wide range of mammalian hosts and a flea vector for transmission, it remains to be elucidated whether YapG has a measurable role in any other stage of plague disease.

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Lenz, Jonathan D., Brenda R. S. Temple, and Virginia L. Miller. "Evolution and Virulence Contributions of the Autotransporter Proteins YapJ and YapK of Yersinia pestis CO92 and Their Homologs in Y. pseudotuberculosis IP32953." Infection and Immunity 80, no. 10 (July 16, 2012): 3693–705. http://dx.doi.org/10.1128/iai.00529-12.

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ABSTRACTYersinia pestis, the causative agent of plague, evolved from the gastrointestinal pathogenYersinia pseudotuberculosis. Both species have numerous type Va autotransporters, most of which appear to be highly conserved. InY. pestisCO92, the autotransporter genesyapKandyapJshare a high level of sequence identity. By comparingyapKandyapJto three homologous genes inY. pseudotuberculosisIP32953 (YPTB0365, YPTB3285, and YPTB3286), we show thatyapKis conserved inY. pseudotuberculosis, whileyapJis unique toY. pestis. All of these autotransporters exhibit >96% identity in the C terminus of the protein and identities ranging from 58 to 72% in their N termini. By extending this analysis to include homologous sequences from numerousY. pestisandY. pseudotuberculosisstrains, we determined that these autotransporters cluster into a YapK (YPTB3285) class and a YapJ (YPTB3286) class. The YPTB3286-like gene of mostY. pestisstrains appears to be inactivated, perhaps in favor of maintainingyapJ. Since autotransporters are important for virulence in many bacterial pathogens, includingY. pestis, any change in autotransporter content should be considered for its impact on virulence. Using established mouse models ofY. pestisinfection, we demonstrated that despite the high level of sequence identity,yapKis distinct fromyapJin its contribution to disseminatedY. pestisinfection. In addition, a mutant lacking both of these genes exhibits an additive attenuation, suggesting nonredundant roles foryapJandyapKin systemicY. pestisinfection. However, the deletion of the homologous genes inY. pseudotuberculosisdoes not seem to impact the virulence of this organism in orogastric or systemic infection models.

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Cotter, Shane E., Neeraj K. Surana, and Joseph W. St. Geme. "Trimeric autotransporters: a distinct subfamily of autotransporter proteins." Trends in Microbiology 13, no. 5 (May 2005): 199–205. http://dx.doi.org/10.1016/j.tim.2005.03.004.

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Sauri, Ana, Zora Soprova, David Wickström, Jan-Willem de Gier, Roel C. Van der Schors, August B. Smit, Wouter S. P. Jong, and Joen Luirink. "The Bam (Omp85) complex is involved in secretion of the autotransporter haemoglobin protease." Microbiology 155, no. 12 (December 1, 2009): 3982–91. http://dx.doi.org/10.1099/mic.0.034991-0.

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Autotransporters are large virulence factors secreted by Gram-negative bacteria. They are synthesized with a C-terminal domain that forms a β-barrel pore in the outer membrane implicated in translocation of the upstream ‘passenger’ domain across the outer membrane. However, recent structural data suggest that the diameter of the β-barrel pore is not sufficient to allow the passage of partly folded structures observed for several autotransporters. Here, we have used a stalled translocation intermediate of the autotransporter Hbp to identify components involved in insertion and translocation of the protein across the outer membrane. At this intermediate stage the β-domain was not inserted and folded as an integral β-barrel in the outer membrane whereas part of the passenger was surface exposed. The intermediate was copurified with the periplasmic chaperone SurA and subunits of the Bam (Omp85) complex that catalyse the insertion and assembly of outer-membrane proteins. The data suggest a critical role for this general machinery in the translocation of autotransporters across the outer membrane.

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Restieri, Concetta, Geneviève Garriss, Marie-Claude Locas, and Charles M. Dozois. "Autotransporter-Encoding Sequences Are Phylogenetically Distributed among Escherichia coli Clinical Isolates and Reference Strains." Applied and Environmental Microbiology 73, no. 5 (January 12, 2007): 1553–62. http://dx.doi.org/10.1128/aem.01542-06.

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ABSTRACT Autotransporters are secreted bacterial proteins exhibiting diverse virulence functions. Various autotransporters have been identified among Escherichia coli associated with intestinal or extraintestinal infections; however, the specific distribution of autotransporter sequences among a diversity of E. coli strains has not been investigated. We have validated the use of a multiplex PCR assay to screen for the presence of autotransporter sequences. Herein, we determined the presence of 13 autotransporter sequences and five allelic variants of antigen 43 (Ag43) among 491 E. coli isolates from human urinary tract infections, diarrheagenic E. coli, and avian pathogenic E. coli (APEC) and E. coli reference strains belonging to the ECOR collection. Clinical isolates were also classified into established phylogenetic groups. The results indicated that Ag43 alleles were significantly associated with clinical isolates (93%) compared to commensal isolates (56%) and that agn43K12 was the most common and widely distributed allele. agn43 allelic variants were also phylogenetically distributed. Sequences encoding espC, espP, and sepA and agn43 alleles EDL933 and RS218 were significantly associated with diarrheagenic E. coli strains compared to other groups. tsh was highly associated with APEC strains, whereas sat was absent from APEC. vat, sat, and pic were associated with urinary tract isolates and were identified predominantly in isolates belonging to either group B2 or D of the phylogenetic groups based on the ECOR strain collection. Overall, the results indicate that specific autotransporter sequences are associated with the source and/or phylogenetic background of strains and suggest that, in some cases, autotransporter gene profiles may be useful for comparative analysis of E. coli strains from clinical, food, and environmental sources.

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Van Ulsen, Peter, Loek Van Alphen, Jan Ten Hove, Floris Fransen, Peter Van Der Ley, and Jan Tommassen. "A Neisserial autotransporter NalP modulating the processing of other autotransporters." Molecular Microbiology 50, no. 3 (October 20, 2003): 1017–30. http://dx.doi.org/10.1046/j.1365-2958.2003.03773.x.

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9

Knudsen, Stine K., Allan Stensballe, Magnus Franzmann, Uffe B. Westergaard, and Daniel E. Otzen. "Effect of glycosylation on the extracellular domain of the Ag43 bacterial autotransporter: enhanced stability and reduced cellular aggregation." Biochemical Journal 412, no. 3 (May 28, 2008): 563–77. http://dx.doi.org/10.1042/bj20071497.

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Autotransporters constitute the biggest group of secreted proteins in Gram-negative bacteria and contain a membrane-bound β-domain and a passenger domain secreted to the extracellular environment via an unusually long N-terminal sequence. Several passenger domains are known to be glycosylated by cytosolic glycosyl transferases, promoting bacterial attachment to mammalian cells. In the present study we describe the effect of glycosylation on the extracellular passenger domain of the Escherichia coli autotransporter Ag43α, which induces frizzy colony morphology and cell settling. We identify 16 glycosylation sites and suggest two possible glycosylation motifs for serine and threonine residues. Glycosylation stabilizes against thermal and chemical denaturation and increases refolding kinetics. Unexpectedly, glycosylation also reduces the stabilizing effect of Ca2+ ions, removes the ability of Ca2+ to promote cell adhesion, reduces the ability of Ag43α-containing cells to form bacterial amyloid and increases the susceptibility of the resulting amyloid to proteolysis. In addition, our results indicate that Ag43α folds without a stable intermediate, unlike pertactin, indicating that autotransporters may arrive at the native state by a variety of different mechanisms despite a common overall structure. A small but significant fraction of Ag43α can survive intact in the periplasm if expressed without the β-domain, suggesting that it is able to adopt a protease-resistant structure prior to translocation across the membrane. The present study demonstrates that glycosylation may play significant roles in structural and functional properties of bacterial autotransporters at many different levels.

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Kostakioti, Maria, and Christos Stathopoulos. "Functional Analysis of the Tsh Autotransporter from an Avian Pathogenic Escherichia coli Strain." Infection and Immunity 72, no. 10 (October 2004): 5548–54. http://dx.doi.org/10.1128/iai.72.10.5548-5554.2004.

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ABSTRACT The temperature-sensitive hemagglutinin (Tsh) is an autotransporter protein secreted by avian-pathogenic Escherichia coli strains that colonize the respiratory tract and lead to airsacculitis, pericarditis, and colisepticemia. It is synthesized as a 140-kDa precursor protein, whose processing results in a 106-kDa passenger domain (Tshs) and a 33-kDa β-domain (Tshβ). The presence of a conserved 7-amino-acid serine protease motif within Tshs classifies the protein in a subfamily of autotransporters, known as serine protease autotransporters of the Enterobacteriaceae. In this study, we report that purified Tshs is capable of adhering to red blood cells, hemoglobin, and the extracellular matrix proteins fibronectin and collagen IV. We also demonstrate that Tshs exerts proteolytic activity against casein, and we provide experimental evidence demonstrating that serine 259 is essential for the protease function. However, this residue is not required for adherence to substrates, and its replacement by an alanine does not abolish binding activity. In summary, our results demonstrate that Tsh is a bifunctional protein with both adhesive and proteolytic properties.

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Hoopman, Todd C., Wei Wang, Chad A. Brautigam, Jennifer L. Sedillo, Thomas J. Reilly, and Eric J. Hansen. "Moraxella catarrhalis Synthesizes an Autotransporter That Is an Acid Phosphatase." Journal of Bacteriology 190, no. 4 (December 7, 2007): 1459–72. http://dx.doi.org/10.1128/jb.01688-07.

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ABSTRACT Moraxella catarrhalis O35E was shown to synthesize a 105-kDa protein that has similarity to both acid phosphatases and autotransporters. The N-terminal portion of the M. catarrhalis acid phosphatase A (MapA) was most similar (the BLAST probability score was 10−10) to bacterial class A nonspecific acid phosphatases. The central region of the MapA protein had similarity to passenger domains of other autotransporter proteins, whereas the C-terminal portion of MapA resembled the translocation domain of conventional autotransporters. Cloning and expression of the M. catarrhalis mapA gene in Escherichia coli confirmed the presence of acid phosphatase activity in the MapA protein. The MapA protein was shown to be localized to the outer membrane of M. catarrhalis and was not detected either in the soluble cytoplasmic fraction from disrupted M. catarrhalis cells or in the spent culture supernatant fluid from M. catarrhalis. Use of the predicted MapA translocation domain in a fusion construct with the passenger domain from another predicted M. catarrhalis autotransporter confirmed the translocation ability of this MapA domain. Inactivation of the mapA gene in M. catarrhalis strain O35E reduced the acid phosphatase activity expressed by this organism, and this mutation could be complemented in trans with the wild-type mapA gene. Nucleotide sequence analysis of the mapA gene from six M. catarrhalis strains showed that this protein was highly conserved among strains of this pathogen. Site-directed mutagenesis of a critical histidine residue (H233A) in the predicted active site of the acid phosphatase domain in MapA eliminated acid phosphatase activity in the recombinant MapA protein. This is the first description of an autotransporter protein that expresses acid phosphatase activity.

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Janakiraman, Anuradha, Kathryn R. Fixen, Andrew N. Gray, Hironori Niki, and Marcia B. Goldberg. "A Genome-Scale Proteomic Screen Identifies a Role for DnaK in Chaperoning of Polar Autotransporters in Shigella." Journal of Bacteriology 191, no. 20 (August 14, 2009): 6300–6311. http://dx.doi.org/10.1128/jb.00833-09.

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ABSTRACT Autotransporters are outer membrane proteins that are widely distributed among gram-negative bacteria. Like other autotransporters, the Shigella autotransporter IcsA, which is required for actin assembly during infection, is secreted at the bacterial pole. In the bacterial cytoplasm, IcsA localizes to poles and potential cell division sites independent of the cell division protein FtsZ. To identify bacterial proteins involved in the targeting of IcsA to the pole in the bacterial cytoplasm, we screened a genome-scale library of E scherichia coli proteins tagged with green fluorescent protein (GFP) for those that displayed a localization pattern similar to that of IcsA-GFP in cells that lack functional FtsZ using a strain carrying a temperature-sensitive ftsZ allele. For each protein that mimicked the localization of IcsA-GFP, we tested whether IcsA localization was dependent on the presence of the protein. Although these approaches did not identify a polar receptor for IcsA, the cytoplasmic chaperone DnaK both mimicked IcsA localization at elevated temperatures as a GFP fusion and was required for the localization of IcsA to the pole in the cytoplasm of E. coli. DnaK was also required for IcsA secretion at the pole in S higella flexneri. The localization of DnaK-GFP to poles and potential cell division sites was dependent on elevated growth temperature and independent of the presence of IcsA or functional FtsZ; native DnaK was found to be enhanced at midcell and the poles. A second Shigella autotransporter, SepA, also required DnaK for secretion, consistent with a role of DnaK more generally in the chaperoning of autotransporter proteins in the bacterial cytoplasm.

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Leo, Jack C., Iwan Grin, and Dirk Linke. "Type V secretion: mechanism(s) of autotransport through the bacterial outer membrane." Philosophical Transactions of the Royal Society B: Biological Sciences 367, no. 1592 (April 19, 2012): 1088–101. http://dx.doi.org/10.1098/rstb.2011.0208.

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Autotransport in Gram-negative bacteria denotes the ability of surface-localized proteins to cross the outer membrane (OM) autonomously. Autotransporters perform this task with the help of a β-barrel transmembrane domain localized in the OM. Different classes of autotransporters have been investigated in detail in recent years; classical monomeric but also trimeric autotransporters comprise many important bacterial virulence factors. So do the two-partner secretion systems, which are a special case as the transported protein resides on a different polypeptide chain than the transporter. Despite the great interest in these proteins, the exact mechanism of the transport process remains elusive. Moreover, different periplasmic and OM factors have been identified that play a role in the translocation, making the term ‘autotransport’ debatable. In this review, we compile the wealth of details known on the mechanism of single autotransporters from different classes and organisms, and put them into a bigger perspective. We also discuss recently discovered or rediscovered classes of autotransporters.

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Kostakioti, Maria, and Christos Stathopoulos. "Role of the α-Helical Linker of the C-Terminal Translocator in the Biogenesis of the Serine Protease Subfamily of Autotransporters." Infection and Immunity 74, no. 9 (September 2006): 4961–69. http://dx.doi.org/10.1128/iai.00103-06.

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ABSTRACT Autotransporters are secreted virulence factors that comprise three domains: an N-terminal signal peptide, an internal passenger domain, and a C-terminal β-domain. The mechanism of passenger translocation across the outer membrane remains undefined, with four models having been proposed: the “hairpin,” the “threading,” the “multimeric,” and the “Omp85 (YaeT)” models. In an attempt to understand autotransporter biogenesis, we screened the sequences of the serine protease subfamily of autotransporters (SPATEs) for conserved features indicative of a common secretion mechanism. Our analyses revealed a strictly conserved 14-amino-acid motif within the predicted α-helical linker region, upstream of the β-domain of SPATEs. We investigated the function of this motif through a mutagenesis approach using Tsh as a model. Our studies demonstrate that mutations throughout the conserved motif do not block insertion of the β-domain into the outer membrane. However, nonconservative mutations of four hydrophobic (V1099, L1102, G1107, and L1109) and three polar (N1100, K1104, and R1105) residues of the motif severely decrease or even abolish Tsh biogenesis. Further studies showed that these mutations interfere with passenger transport across the outer membrane. Bioinformatical analyses suggest that the critical polar and hydrophobic amino acids localize on opposite sides of the helix that runs through the β-barrel pore. Our data indicate that the conserved motif is important for passenger secretion across the outer membrane and that mutations in certain residues severely affect the secretion process. We discuss how these results fit with the four proposed models for autotransporter secretion and potential applications in antimicrobial and vaccine development.

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Al-Hasani, Keith, Ian R. Henderson, Harry Sakellaris, Kumar Rajakumar, Travis Grant, James P. Nataro, Roy Robins-Browne, and Ben Adler. "The sigA Gene Which Is Borne on the shePathogenicity Island of Shigella flexneri 2a Encodes an Exported Cytopathic Protease Involved in Intestinal Fluid Accumulation." Infection and Immunity 68, no. 5 (May 1, 2000): 2457–63. http://dx.doi.org/10.1128/iai.68.5.2457-2463.2000.

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ABSTRACT In this study, the sigA gene situated on theshe pathogenicity island of Shigella flexneri2a was cloned and characterized. Sequence analysis showed thatsigA encodes a 139.6-kDa protein which belongs to the SPATE (serine protease autotransporters of Enterobacteriaceae) subfamily of autotransporter proteins. The demonstration that SigA is autonomously secreted from the cell to yield a 103-kDa processed form and possesses a conserved C-terminal domain for export from the cell were consistent with the autotransporter pathway of secretion. Functional analysis showed that SigA is a secreted temperature-regulated serine protease capable of degrading casein. SigA was cytopathic for HEp-2 cells, suggesting that it may be a cell-altering toxin with a role in the pathogenesis ofShigella infections. SigA was at least partly responsible for the ability of S. flexneri to stimulate fluid accumulation in ligated rabbit ileal loops.

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Pina, Sonia, Alex Olvera, Anna Barceló, and Albert Bensaid. "Trimeric Autotransporters of Haemophilus parasuis: Generation of an Extensive Passenger Domain Repertoire Specific for Pathogenic Strains." Journal of Bacteriology 191, no. 2 (November 14, 2008): 576–87. http://dx.doi.org/10.1128/jb.00703-08.

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ABSTRACT Haemophilus parasuis is the agent responsible for causing Glässer's disease, but little is known about the pathogenic determinants of this major pig disease. Here we describe, for the pathogenic strain Nagasaki, the molecular characterization of 13 trimeric autotransporters as assessed by the presence of YadA C-terminal translocator domains which were classified into three groups. All passenger domains possess motifs and repeats characteristic of adhesins, hemagglutinins, and invasins with various centrally located copies of collagen-like repeats. This domain architecture is shared with two trimeric autotransporter proteins of H. somnus 129Pt. Genomic comparison by microarray hybridization demonstrated homologies among H. parasuis virulent strains and high divergence with respect to nonvirulent strains. Therefore, these genes were named vtaA (virulence-associated trimeric autotransporters). The sequencing of 17 homologous vtaA genes of different invasive strains highlighted an extensive mosaic structure. Based also on the presence of DNA uptake signal sequences within the vtaA genes, we propose a mechanism of evolution by which gene duplication and the accumulation of mutations and recombinations, plus the lateral gene transfer of the passenger domain, led to the diversity of this multigene family. This study provides insights to help understand the tissue colonization and invasiveness characteristic of H. parasuis pathogenic strains.

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Veiga, Esteban, Víctor de Lorenzo, and Luis Angel Fernández. "Autotransporters as Scaffolds for Novel Bacterial Adhesins: Surface Properties of Escherichia coli Cells Displaying Jun/Fos Dimerization Domains." Journal of Bacteriology 185, no. 18 (September 15, 2003): 5585–90. http://dx.doi.org/10.1128/jb.185.18.5585-5590.2003.

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ABSTRACT Hybrid proteins containing the β-autotransporter domain of the immunoglobulin A (IgA) protease of Neisseria gonorrhoea (IgAβ) and the partner leucine zippers of the eukaryotic transcriptional factors Fos and Jun were expressed in Escherichia coli. Such fusion proteins targeted the leucine zipper modules to the cell surface. Cells displaying the Junβ sequence flocculated shortly after induction of the hybrid protein. E. coli cells expressing separately Fosβ and Junβ chimeras formed stable bacterial consortia. These associations were physically held by tight intercell ties caused by the protein-protein interactions of matching dimerization domains. The role of autotransporters in the emergence of new adhesins is discussed.

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Benz, Inga, Tessa van Alen, Julia Bolte, Mirka E. Wörmann, and M. Alexander Schmidt. "Modulation of transcription and characterization of the promoter organization of the autotransporter adhesin heptosyltransferase and the autotransporter adhesin AIDA-I." Microbiology 156, no. 4 (April 1, 2010): 1155–66. http://dx.doi.org/10.1099/mic.0.032292-0.

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In Gram-negative bacteria, autotransporter proteins constitute the largest family of secreted proteins, and exhibit many different functions. In recent years, research has largely focused on mechanisms of autotransporter protein translocation, where several alternative models are still being discussed. In contrast, the biogenesis of only a few autotransporters has been studied and, likewise, regulation of expression has received only very limited attention. The glycosylated autotransporter adhesin involved in diffuse adherence (AIDA)-I system consists of the aah gene, encoding a specific autotransporter adhesin heptosyltransferase (AAH), and the aidA gene, encoding the autotransporter protein (AIDA-I). In this study, we investigated the promoter organization and transcription of these two genes using reporter plasmids carrying lacZ transcriptional fusions. The two genes, aah and aidA, are transcribed as a bicistronic message. However, aidA is additionally transcribed from its own promoter. There are two distinct start sites for each of the two genes. Interestingly, transcription of both genes is enhanced in hns and rfaH mutant backgrounds. Furthermore, we addressed the influence of environmental factors and different genetic backgrounds of Escherichia coli K-12 strains on transcription activity. We found that transcription varied considerably in different E. coli K-12 laboratory strains and under different growth conditions.

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Saunders, Jane. "Autotransporters — monomers and multimers." Nature Reviews Microbiology 2, no. 5 (May 2004): 354. http://dx.doi.org/10.1038/nrmicro894.

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Litwin, Christine M., and Joel M. Johnson. "Identification, Cloning, and Expression of the CAMP-Like Factor Autotransporter Gene (cfa) of Bartonella henselae." Infection and Immunity 73, no. 7 (July 2005): 4205–13. http://dx.doi.org/10.1128/iai.73.7.4205-4213.2005.

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ABSTRACT The CAMP reaction was first described by Christie et al. (R. Christie, N. E. Atkins, and E. Munch-Petersen, Aust. J. Exp. Biol. 22:197-200, 1944) as the synergistic lysis of sheep red blood cells by Staphylococcus aureus sphingomyelinase and CAMP factor (cohemolysin), a secreted protein from group B streptococci. We observed a CAMP-like reaction when Bartonella henselae was grown in close proximity to S. aureus on 5% sheep blood agar. This study describes the cloning, sequencing, and characterization of a CAMP-like factor autotransporter gene (cfa) from B. henselae. A cosmid library of B. henselae ATCC 49793 was constructed using SuperCos1 in Escherichia coli XL1-Blue MR. Cosmids were screened for the CAMP reaction, and a quantitative cohemolysis microtiter assay was developed using purified sphingomyelinase. Cosmid clones with the strongest cohemolytic reaction had similar restriction enzyme patterns. A DNA fragment that expressed the cohemolysin determinant was subcloned in a 7,200-bp StuI-BamHI fragment which contained a 6,024-bp open reading frame. The deduced amino acid sequence showed homology to the family of autotransporters. The autotransporters are a group of proteins that mediate their own export through the outer membrane. They contain an N-terminal passenger region, the α-domain, and a C-terminal transporter region, the β-domain. The α-domain contained four, nearly identical 42-amino-acid repeats and showed homology to the family of RTX (repeat in toxin) hemolysins. The concentrated supernatant of the recombinant strain expressed a protein with a molecular mass of 180 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis consistent with the calculated molecular weight of the secreted α-domain. In conclusion, we have characterized a novel secreted cohemolysin autotransporter protein of B. henselae.

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Lattemann, Claus T., Jochen Maurer, Elke Gerland, and Thomas F. Meyer. "Autodisplay: Functional Display of Active β-Lactamase on the Surface of Escherichia coli by the AIDA-I Autotransporter." Journal of Bacteriology 182, no. 13 (July 1, 2000): 3726–33. http://dx.doi.org/10.1128/jb.182.13.3726-3733.2000.

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ABSTRACT Members of the protein family of immunoglobulin A1 protease-like autotransporters comprise multidomain precursors consisting of a C-terminal autotransporter domain that promotes the translocation of N-terminally attached passenger domains across the cell envelopes of gram-negative bacteria. Several autotransporter domains have recently been shown to efficiently promote the export of heterologous passenger domains, opening up an effective tool for surface display of heterologous proteins. Here we report on the autotransporter domain of the Escherichia coli adhesin involved in diffuse adherence (AIDA-I), which was genetically fused to the C terminus of the periplasmic enzyme β-lactamase, leading to efficient expression of the fusion protein in E. coli. The β-lactamase moiety of the fusion protein was presented on the bacterial surface in a stable manner, and the surface-located β-lactamase was shown to be enzymatically active. Enzymatic activity was completely removed by protease treatment, indicating that surface display of β-lactamase was almost quantitative. The periplasmic domain of the outer membrane protein OmpA was not affected by externally added proteases, demonstrating that the outer membranes of E. coli cells expressing the β-lactamase AIDA-I fusion protein remained physiologically intact.

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Ruiz-Perez, Fernando, Ian R. Henderson, Denisse L. Leyton, Amanda E. Rossiter, Yinghua Zhang, and James P. Nataro. "Roles of Periplasmic Chaperone Proteins in the Biogenesis of Serine Protease Autotransporters of Enterobacteriaceae." Journal of Bacteriology 191, no. 21 (September 4, 2009): 6571–83. http://dx.doi.org/10.1128/jb.00754-09.

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ABSTRACT The serine protease autotransporters of Enterobacteriaceae (SPATEs) represent a large family of virulence factors. The prevailing model for autotransporter secretion comprises entry to the periplasm via the Sec apparatus, followed by an obscure series of steps in which the C terminus of the periplasmic species inserts into the outer membrane as a β-barrel protein, accompanied by translocation of the passenger domain to the bacterial cell surface. Little is known about the fate of the autotransporter proteins in the periplasm, including whether accessory periplasmic proteins are involved in translocation to the external milieu. Here we studied the role of the major periplasmic chaperones in the biogenesis of EspP, a prototype SPATE protein produced by Escherichia coli O157:H7. The yeast two-hybrid approach, secretion analysis of chaperone mutant strains, and surface plasmon resonance analysis (SPR) revealed direct protein-protein interactions between the periplasmic SurA and DegP chaperones and either the EspP-β or EspP passenger domains. The secretion of EspP was moderately reduced in the surA and skp mutant strains but severely impaired in the degP background. Site-directed mutagenesis of highly conserved aromatic amino acid residues in the SPATE family resulted in ∼80% reduction of EspP secretion. Synthetic peptides containing aromatic residues derived from the EspP passenger domain blocked DegP and SurA binding to the passenger domain. SPR suggested direct protein-protein interaction between periplasmic chaperones and the unfolded EspP passenger domain. Our data suggest that translocation of AT proteins may require accessory factors, calling into question the moniker “autotransporter.”

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Litwin, Christine M., Mindy L. Rawlins, and Erica M. Swenson. "Characterization of an Immunogenic Outer Membrane Autotransporter Protein, Arp, of Bartonella henselae." Infection and Immunity 75, no. 11 (September 4, 2007): 5255–63. http://dx.doi.org/10.1128/iai.00533-07.

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ABSTRACT Bartonella henselae is a recently recognized pathogenic bacterium associated with cat scratch disease, bacillary angiomatosis, and bacillary peliosis. This study describes the cloning, sequencing, and characterization of an antigenic autotransporter gene from B. henselae. A cloned 6.0-kb BclI-EcoRI DNA fragment expresses a 120-kDa B. henselae protein immunoreactive with 21.2% of sera from patients positive for B. henselae immunoglobulin G antibodies by indirect immunofluorescence, with 97.3% specificity and no cross-reactivity with antibodies against various other organisms. DNA sequencing of the clone revealed one open reading frame of 4,320 bp with a deduced amino acid sequence that shows homology to the family of autotransporters. The autotransporters are a group of proteins that mediate their own export through the outer membrane and consist of a passenger region, the α-domain, and an outer membrane transporter region, the β-domain. The passenger domain shows homology to a family of pertactin-like adhesion proteins and contains seven, nearly identical 48-amino-acid repeats not found in any other bacterial or Bartonella DNA sequences. The passenger α-domain has a calculated molecular mass of 117 kDa, and the transporter β-domain has a calculated molecular mass of 36 kDa. The clone expresses a 120-kDa protein and a protein that migrates at approximately 38 kDa exclusively in the outer membrane protein fraction, suggesting that the 120-kDa passenger protein remains associated with the outer membrane after cleavage from the 36-kDa transporter.

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Bernstein, Harris D. "Are bacterial ‘autotransporters’ really transporters?" Trends in Microbiology 15, no. 10 (October 2007): 441–47. http://dx.doi.org/10.1016/j.tim.2007.09.007.

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Pokharel, Pravil, Hajer Habouria, Hicham Bessaiah, and Charles M. Dozois. "Serine Protease Autotransporters of the Enterobacteriaceae (SPATEs): Out and About and Chopping It Up." Microorganisms 7, no. 12 (November 21, 2019): 594. http://dx.doi.org/10.3390/microorganisms7120594.

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Autotransporters are secreted proteins with multiple functions produced by a variety of Gram-negative bacteria. In Enterobacteriaceae, a subgroup of these autotransporters are the SPATEs (serine protease autotransporters of Enterobacteriaceae). SPATEs play a crucial role in survival and virulence of pathogens such as Escherichia coli and Shigella spp. and contribute to intestinal and extra-intestinal infections. These high molecular weight proteases are transported to the external milieu by the type Va secretion system and function as proteases with diverse substrate specificities and biological functions including adherence and cytotoxicity. Herein, we provide an overview of SPATEs and discuss recent findings on the biological roles of these secreted proteins, including proteolysis of substrates, adherence to cells, modulation of the immune response, and virulence in host models. In closing, we highlight recent insights into the regulation of expression of SPATEs that could be exploited to understand fundamental SPATE biology.

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Cotter, Shane E., Neeraj K. Surana, Susan Grass, and Joseph W. St. Geme. "Trimeric Autotransporters Require Trimerization of the Passenger Domain for Stability and Adhesive Activity." Journal of Bacteriology 188, no. 15 (August 1, 2006): 5400–5407. http://dx.doi.org/10.1128/jb.00164-06.

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ABSTRACT In recent years, structural studies have identified a number of bacterial, viral, and eukaryotic adhesive proteins that have a trimeric architecture. The prototype examples in bacteria are the Haemophilus influenzae Hia adhesin and the Yersinia enterocolitica YadA adhesin. Both Hia and YadA are members of the trimeric-autotransporter subfamily and are characterized by an internal passenger domain that harbors adhesive activity and a short C-terminal translocator domain that inserts into the outer membrane and facilitates delivery of the passenger domain to the bacterial surface. In this study, we examined the relationship between trimerization of the Hia and YadA passenger domains and the capacity for adhesive activity. We found that subunit-subunit interactions and stable trimerization are essential for native folding and stability and ultimately for full-level adhesive activity. These results raise the possibility that disruption of the trimeric architecture of trimeric autotransporters, and possibly other trimeric adhesins, may be an effective strategy to eliminate adhesive activity.

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Tame, Jeremy R. H. "Autotransporter protein secretion." BioMolecular Concepts 2, no. 6 (December 1, 2011): 525–36. http://dx.doi.org/10.1515/bmc.2011.045.

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AbstractAutotransporter proteins are a large family of virulence factors secreted from Gram-negative bacteria by a unique mechanism. First described in the 1980s, these proteins have a C-terminal region that folds into a β-barrel in the bacterial outer membrane. The so-called passenger domain attached to this barrel projects away from the cell surface and may be liberated from the cell by self-cleavage or surface proteases. Although the majority of passenger domains have a similar β-helical structure, they carry a variety of subdomains, allowing them to carry out widely differing functions related to pathogenesis. Considerable biochemical and structural characterisation of the barrel domain has shown that ‘autotransporters’ in fact require a conserved and essential protein complex in the outer membrane for correct folding. Although the globular domains of this complex projecting into the periplasmic space have also been structurally characterised, the overall secretion pathway of the autotransporters remains highly puzzling. It was presumed for many years that the passenger domain passed through the centre of the barrel domain to reach the cell surface, driven at least in part by folding. This picture is complicated by conflicting data, and there is currently little hard information on the true nature of the secretion intermediates. As well as their medical importance therefore, autotransporters are proving to be an excellent system to study the folding and membrane insertion of outer membrane proteins in general. This review focuses on structural aspects of autotransporters; their many functions in pathogenesis are beyond its scope.

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Müller, Daniel, Inga Benz, Damini Tapadar, Christian Buddenborg, Lilo Greune, and M. Alexander Schmidt. "Arrangement of the Translocator of the Autotransporter Adhesin Involved in Diffuse Adherence on the Bacterial Surface." Infection and Immunity 73, no. 7 (July 2005): 3851–59. http://dx.doi.org/10.1128/iai.73.7.3851-3859.2005.

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ABSTRACT Autotransporters of gram-negative bacteria are single-peptide secretion systems that consist of a functional N-terminal α-domain (“passenger”) fused to a C-terminal β-domain (“translocator”). How passenger proteins are translocated through the outer membrane has not been resolved, and at present essentially three different models are discussed. In the widely accepted “hairpin model” the passenger proteins are translocated through a channel formed by the β-barrel of the translocator that is integrated in the outer membrane. This model has been challenged by a recent proposal for a general autotransporter model suggesting that there is a hexameric translocation pore that is generated by the oligomerization of six β-domains. A third model suggests that conserved Omp85 participates in autotransporter integration and passenger protein translocation. To examine these models, in this study we investigated the presence of putative oligomeric structures of the translocator of the autotransporter adhesin involved in diffuse adherence (AIDA) in vivo by cross-linking techniques. Furthermore, the capacity of isolated AIDA fusion proteins to form oligomers was studied in vitro by several complementary analytical techniques, such as analytical gel filtration, electron microscopy, immunogold labeling, and cross-linking of recombinant autotransporter proteins in which different passenger proteins were fused to the AIDA translocator. Our results show that the AIDA translocator is mostly present as a monomer. Only a fraction of the AIDA autotransporter was found to form dimers on the bacterial surface and in solution. Higher-order structures, such as hexamers, were not detected either in vivo or in vitro and can therefore be excluded as functional moieties for the AIDA autotransporter.

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Letley, Darren P., Joanne L. Rhead, Keith Bishop, and John C. Atherton. "Paired cysteine residues are required for high levels of the Helicobacter pylori autotransporter VacA." Microbiology 152, no. 5 (May 1, 2006): 1319–25. http://dx.doi.org/10.1099/mic.0.28548-0.

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The Helicobacter pylori vacuolating cytotoxin VacA shares homology in its C-terminal domain with many autotransporter proteins, suggesting a similar mechanism of secretion. Like most autotransporters, VacA contains a single pair of cysteine residues located near the C-terminus of the passenger domain. This study aimed to investigate the role of these conserved cysteine residues. This involved changing each cysteine in the VacA passenger domain to serine, quantifying the effect on VacA levels and assessing toxin activity in H. pylori. It was shown that both cysteine residues were required for high VacA levels, although mutation of each cysteine reduced toxin amounts to differing extents, implying that their importance was not simply for intramolecular disulphide bond formation. Although less VacA was observed for the cysteine mutants, vacuolating activity was detected, showing that the cysteines were not required for VacA function.

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Pokharel, Pravil, Juan Manuel Díaz, Hicham Bessaiah, Sébastien Houle, Alma Lilián Guerrero-Barrera, and Charles M. Dozois. "The Serine Protease Autotransporters TagB, TagC, and Sha from Extraintestinal Pathogenic Escherichia coli Are Internalized by Human Bladder Epithelial Cells and Cause Actin Cytoskeletal Disruption." International Journal of Molecular Sciences 21, no. 9 (April 26, 2020): 3047. http://dx.doi.org/10.3390/ijms21093047.

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TagB, TagC (tandem autotransporter genes B and C), and Sha (Serine-protease hemagglutinin autotransporter) are recently described members of the SPATE (serine protease autotransporters of Enterobacteriaceae) family. These SPATEs can cause cytopathic effects on bladder cells and contribute to urinary tract infection in a mouse model. Bladder epithelial cells form an important barrier in the urinary tract. Some SPATEs produced by pathogenic E. coli are known to breach the bladder epithelium. The capacity of these newly described SPATEs to alter bladder epithelial cells and the role of the serine protease active site were investigated. All three SPATE proteins were internalized by bladder epithelial cells and altered the distribution of actin cytoskeleton. Sha and TagC were also shown to degrade mucin and gelatin respectively. Inactivation of the serine catalytic site in each of these SPATEs did not affect secretion of the SPATEs from bacterial cells, but abrogated entry into epithelial cells, cytotoxicity, and proteolytic activity. Thus, our results show that the serine catalytic triad of these proteins is required for internalization in host cells, actin disruption, and degradation of host substrates such as mucin and gelatin.

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Parreira, V. R., and C. L. Gyles. "A Novel Pathogenicity Island Integrated Adjacent to the thrW tRNA Gene of Avian Pathogenic Escherichia coli Encodes a Vacuolating Autotransporter Toxin." Infection and Immunity 71, no. 9 (September 2003): 5087–96. http://dx.doi.org/10.1128/iai.71.9.5087-5096.2003.

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ABSTRACT We report the complete nucleotide sequence and genetic organization of the Vat-encoding pathogenicity island (PAI) of avian pathogenic Escherichia coli strain Ec222. The 22,139-bp PAI is situated adjacent to the 3′ terminus of the thrW tRNA gene, has a G+C content of 41.2%, and includes a bacteriophage SfII integrase gene, mobile genetic elements, two open reading frames with products exhibiting sequence similarity to known proteins, and several other open reading frames of unknown function. The PAI encodes an autotransporter protein, Vat (vacuolating autotransporter toxin), which induces the formation of intracellular vacuoles resulting in cytotoxic effects similar to those caused by the VacA toxin from Helicobacter pylori. The predicted 148.3-kDa protein product possesses the three domains that are typical of serine protease autotransporters of Enterobacteriaceae: an N-terminal signal sequence of 55 amino acids, a 111.8-kDa passenger domain containing a modified serine protease site (ATSGSG), and a C-terminal outer membrane translocator of 30.5 kDa. Vat has 75% protein homology with the hemagglutinin Tsh, an autotransporter of avian pathogenic E. coli. A vat deletion mutant of Ec222 showed no virulence in respiratory and cellulitis infection models of disease in broiler chickens. We conclude that the newly described PAI and Vat may be involved in the pathogenicity of avian septicemic E. coli strain Ec222 and other avian pathogenic E. coli strains.

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Hiss, Jan A., and Gisbert Schneider. "Domain Organization of Long Autotransporter Signal Sequences." Bioinformatics and Biology Insights 3 (January 2009): BBI.S3411. http://dx.doi.org/10.4137/bbi.s3411.

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Bacterial autotransporters represent a diverse family of proteins that autonomously translocate across the inner membrane of Gram-negative bacteria via the Sec complex and across the outer bacterial membrane. They often possess exceptionally long N-terminal signal sequences. We analyzed 90 long signal sequences of bacterial autotransporters and members of the two-partner secretion pathway in silico and describe common domain organization found in 79 of these sequences. The domains are in agreement with previously published experimental data. Our algorithmic approach allows for the systematic identification of functionally different domains in long signal sequences.

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Navarro-Garcia, Fernando, and Waldir P. Elias. "Autotransporters and virulence of enteroaggregativeE. coli." Gut Microbes 2, no. 1 (January 2011): 13–24. http://dx.doi.org/10.4161/gmic.2.1.14933.

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Ali, Tehmeena, Neil J. Oldfield, Karl G. Wooldridge, David P. Turner, and Dlawer A. A. Ala'Aldeen. "Functional Characterization of AasP, a Maturation Protease Autotransporter Protein of Actinobacillus pleuropneumoniae." Infection and Immunity 76, no. 12 (October 13, 2008): 5608–14. http://dx.doi.org/10.1128/iai.00085-08.

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ABSTRACT Actinobacillus pleuropneumoniae is the etiological agent of porcine pleuropneumonia, a highly contagious respiratory infection in pigs. AasP, a putative subtilisin-like serine protease autotransporter, has recently been identified in A. pleuropneumoniae. We hypothesized that, similarly to other autotransporters of this type, AasP may undergo autocatalytic cleavage resulting in release of the passenger domain of the protein. Furthermore, AasP may be responsible for cleavage of other A. pleuropneumoniae outer membrane proteins. To address these hypotheses, the aasP gene was cloned and the expressed recombinant AasP protein used to raise monospecific rabbit antiserum. Immunoblot analysis of whole-cell lysates and secreted proteins demonstrated that AasP does not undergo proteolytic cleavage. Immunoblot analysis also confirmed that AasP is universally expressed by A. pleuropneumoniae. Confirmation of the maturation protease function of AasP was obtained through phenotypic analysis of an A. pleuropneumoniae aasP deletion mutant and by functional complementation. Comparison of the secreted proteins of the wild type, an aasP mutant derivative, and an aasP mutant complemented in trans led to the identification of OmlA protein fragments that were present only in the secreted-protein preparations of the wild-type and complemented strains, indicating that AasP is involved in modification of OmlA. This is the first demonstration of a function for any autotransporter protein in Actinobacillus pleuropneumoniae.

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Roussel-Jazédé, Virginie, Ilse Jongerius, Martine P. Bos, Jan Tommassen, and Peter van Ulsen. "NalP-Mediated Proteolytic Release of Lactoferrin-Binding Protein B from the Meningococcal Cell Surface." Infection and Immunity 78, no. 7 (April 26, 2010): 3083–89. http://dx.doi.org/10.1128/iai.01193-09.

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ABSTRACT Bacteria have developed several mechanisms for iron uptake during colonization of mammalian hosts, where the availability of free iron is limiting for growth. Neisseria meningitidis expresses under iron-limiting conditions a receptor complex consisting of the lactoferrin-binding proteins A (LbpA) and LbpB to acquire iron from lactoferrin, which is abundantly present on the mucosal surfaces of the human nasopharynx. LbpA is an integral outer membrane-embedded iron transporter, whereas LbpB is a cell surface-exposed lipoprotein. In this study, we demonstrate that LbpB is also released into the culture medium. We identified NalP, an autotransporter known to be involved in the processing of other autotransporters, as the protease responsible for LbpB release. This release of LbpB reduced the complement-mediated killing of the bacteria when incubated with an LbpB-specific bactericidal antiserum. Since antibodies directed against LbpB are found in convalescent-patient sera, the release of an immunogenic protein as LbpB may represent a novel means for N. meningitidis to escape the human immune response.

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Henderson, Ian R., John Czeczulin, Carlos Eslava, Fernando Noriega, and James P. Nataro. "Characterization of Pic, a Secreted Protease ofShigella flexneri and EnteroaggregativeEscherichia coli." Infection and Immunity 67, no. 11 (November 1, 1999): 5587–96. http://dx.doi.org/10.1128/iai.67.11.5587-5596.1999.

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ABSTRACT We have identified and characterized a secreted protein, designated Pic, which is encoded on the chromosomes of enteroaggregativeEscherichia coli (EAEC) 042 and Shigella flexneri 2457T. The product of the pic gene is synthesized as a 146.5-kDa precursor molecule which is processed at the N and C termini during secretion, allowing the release of a mature protein (109.8 kDa) into the culture supernatant. The deduced amino acid sequence of Pic shows high homology to autotransporter proteins, particularly a subgroup termed the SPATEs (serine protease autotransporters of the Enterobacteriaceae). Present in all members of this subgroup is a motif similar to the active sites of certain serine proteases. Pic catalyzes gelatin degradation, which can be abolished by disruption of the predicted proteolytic active site. Functional analysis of the Pic protein implicates this factor in mucinase activity, serum resistance, and hemagglutination. Our data suggest that Pic may be a multifunctional protein involved in enteric pathogenesis.

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Bialer, Magalí G., Gabriela Sycz, Florencia Muñoz González, Mariana C. Ferrero, Pablo C. Baldi, and Angeles Zorreguieta. "Adhesins of Brucella: Their Roles in the Interaction with the Host." Pathogens 9, no. 11 (November 12, 2020): 942. http://dx.doi.org/10.3390/pathogens9110942.

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A central aspect of Brucella pathogenicity is its ability to invade, survive, and replicate in diverse phagocytic and non-phagocytic cell types, leading to chronic infections and chronic inflammatory phenomena. Adhesion to the target cell is a critical first step in the invasion process. Several Brucella adhesins have been shown to mediate adhesion to cells, extracellular matrix components (ECM), or both. These include the sialic acid-binding proteins SP29 and SP41 (binding to erythrocytes and epithelial cells, respectively), the BigA and BigB proteins that contain an Ig-like domain (binding to cell adhesion molecules in epithelial cells), the monomeric autotransporters BmaA, BmaB, and BmaC (binding to ECM components, epithelial cells, osteoblasts, synoviocytes, and trophoblasts), the trimeric autotransporters BtaE and BtaF (binding to ECM components and epithelial cells) and Bp26 (binding to ECM components). An in vivo role has also been shown for the trimeric autotransporters, as deletion mutants display decreased colonization after oral and/or respiratory infection in mice, and it has also been suggested for BigA and BigB. Several adhesins have shown unipolar localization, suggesting that Brucella would express an adhesive pole. Adhesin-based vaccines may be useful to prevent brucellosis, as intranasal immunization in mice with BtaF conferred high levels of protection against oral challenge with B. suis.

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Patel, Seema K., Jimmie Dotson, Kenneth P. Allen, and James M. Fleckenstein. "Identification and Molecular Characterization of EatA, an Autotransporter Protein of Enterotoxigenic Escherichia coli." Infection and Immunity 72, no. 3 (March 2004): 1786–94. http://dx.doi.org/10.1128/iai.72.3.1786-1794.2004.

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ABSTRACT Enterotoxigenic Escherichia coli (ETEC) strains remain a formidable cause of diarrheal disease. To identify novel surface proteins of ETEC, we performed TnphoA mutagenesis of prototype ETEC strain H10407 and discovered a secreted protein not previously recognized in ETEC. DNA sequencing of the interrupted locus in mutant TnphoA.977 revealed a candidate 4,095-bp open reading frame without significant homology to commensal E. coli K-12 genomic DNA. Translation of this sequence revealed that it encoded a predicted peptide of 147.7 kDa that bears significant homology to members of the autotransporter family of bacterial virulence factors, particularly the serine protease autotransporters of the Enterobacteriaceae proteins. The gene identified in H10407, eatA (ETEC autotransporter A), encodes a potential serine protease motif (GDSGSP) in the secreted amino-terminal domain, and the predicted peptide shows more than 80% homology with SepA, a virulence protein secreted by Shigella flexneri. DNA hybridization and PCR demonstrated that eatA resides on the 92-kDa pCS1 virulence plasmid of H10407 and that it is present in multiple clinical ETEC strains. Immunoblots with antisera directed against a recombinant EatA passenger protein fragment identified a 110-kDa protein in supernatants purified from H10407 but not from the TnphoA.977 mutant or H10407-P, which lacks pCS1. EatA possesses serine protease activity that is abolished by mutations within a serine protease catalytic triad formed by residues H134, D162, and S267. Finally, interruption of the eatA gene retarded fluid accumulation in the rabbit ileal loop model, suggesting that this autotransporter contributes to the virulence of ETEC.

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Campos, Cristine G., Matthew S. Byrd, and Peggy A. Cotter. "Functional Characterization of Burkholderia pseudomallei Trimeric Autotransporters." Infection and Immunity 81, no. 8 (May 28, 2013): 2788–99. http://dx.doi.org/10.1128/iai.00526-13.

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ABSTRACTBurkholderia pseudomalleiis a tier 1 select agent and the causative agent of melioidosis, a severe and often fatal disease with symptoms ranging from acute pneumonia and septic shock to a chronic infection characterized by abscess formation in the lungs, liver, and spleen. Autotransporters (ATs) are exoproteins belonging to the type V secretion system family, with many playing roles in pathogenesis. The genome ofB. pseudomalleistrain 1026b encodes nine putative trimeric AT proteins, of which only four have been described. Using a bioinformatic approach, we annotated putative domains within each trimeric AT protein, excluding the well-studied BimA protein, and found short repeated sequences unique toBurkholderiaspecies, as well as an unexpectedly large proportion of ATs with extended signal peptide regions (ESPRs). To characterize the role of trimeric ATs in pathogenesis, we constructed disruption or deletion mutations in each of eight AT-encoding genes and evaluated the resulting strains for adherence to, invasion of, and plaque formation in A549 cells. The majority of the ATs (and/or the proteins encoded downstream) contributed to adherence to and efficient invasion of A549 cells. Using a BALB/c mouse model of infection, we determined the contributions of each AT to bacterial burdens in the lungs, liver, and spleen. At 48 h postinoculation, only one strain, Bp340::pDbpaC, demonstrated a defect in dissemination and/or survival in the liver, indicating that BpaC is required for wild-type virulence in this model.

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Heimer, Susan R., David A. Rasko, C. Virginia Lockatell, David E. Johnson, and Harry L. T. Mobley. "Autotransporter Genes pic and tsh Are Associated with Escherichia coli Strains That Cause Acute Pyelonephritis and Are Expressed during Urinary Tract Infection." Infection and Immunity 72, no. 1 (January 2004): 593–97. http://dx.doi.org/10.1128/iai.72.1.593-597.2004.

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ABSTRACT We have identified two chromosomal open reading frames in uropathogenic Escherichia coli (UPEC) strain CFT073 which are highly homologous to serine protease autotransporters Pic and Tsh. Both cloned determinants were correlated with the presence of 105- to 110-kDa proteins in the culture supernatants. Furthermore, in cellular fractionation experiments, 30-kDa polypeptides were identified in the outer membrane; we speculated that these proteins are the β-barrel portions of the autotransporter homologues. Furthermore, Pic-containing culture supernatants have serine protease activity. In reverse transcription-PCR analyses, the expression of the pic and tsh genes in E. coli CFT073 was higher in broth cultures grown at 37°C than at 25°C. Moreover, pic and tsh were expressed by bacteria isolated from urine of transurethrally infected mice. The tsh determinant was identified in 63% of our clinical UPEC strain isolates (n = 87) and in 33% of fecal strains (n = 27), whereas pic was present in 31% of the pyelonephritis (n = 67) and 7% of the fecal strains. There was no significant correlation between cystitis strains (n = 20) and the pic determinant.

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Li, Ganwu, Yaping Feng, Subhashinie Kariyawasam, Kelly A. Tivendale, Yvonne Wannemuehler, Fanghong Zhou, Catherine M. Logue, Cathy L. Miller, and Lisa K. Nolan. "AatA Is a Novel Autotransporter and Virulence Factor of Avian Pathogenic Escherichia coli." Infection and Immunity 78, no. 3 (December 22, 2009): 898–906. http://dx.doi.org/10.1128/iai.00513-09.

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ABSTRACT Autotransporters (AT) are widespread in Gram-negative bacteria, and many of them are involved in virulence. An open reading frame (APECO1_O1CoBM96) encoding a novel AT was located in the pathogenicity island of avian pathogenic Escherichia coli (APEC) O1's virulence plasmid, pAPEC-O1-ColBM. This 3.5-kb APEC autotransporter gene (aatA) is predicted to encode a 123.7-kDa protein with a 25-amino-acid signal peptide, an 857-amino-acid passenger domain, and a 284-amino-acid β domain. The three-dimensional structure of AatA was also predicted by the threading method using the I-TASSER online server and then was refined using four-body contact potentials. Molecular analysis of AatA revealed that it is translocated to the cell surface, where it elicits antibody production in infected chickens. Gene prevalence analysis indicated that aatA is strongly associated with E. coli from avian sources but not with E. coli isolated from human hosts. Also, AatA was shown to enhance adhesion of APEC to chicken embryo fibroblast cells and to contribute to APEC virulence.

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Oliver, David C., George Huang, and Rachel C. Fernandez. "Identification of Secretion Determinants of the Bordetella pertussis BrkA Autotransporter." Journal of Bacteriology 185, no. 2 (January 15, 2003): 489–95. http://dx.doi.org/10.1128/jb.185.2.489-495.2003.

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ABSTRACT The autotransporters comprise a functionally diverse family of gram-negative proteins that mediate their own export across the bacterial outer membrane. They consist of an amino-terminal passenger region called the “α-domain” and the structural hallmark of the autotransporter family, a carboxy-terminal transporter region usually referred to as the “β-domain.” The passenger region can be quite diverse and constitutes the effector functions of these proteins, whereas the C-terminal region is conserved and is responsible for translocating the passenger moiety across the outer membrane. BrkA is the 103-kDa autotransporter protein in Bordetella pertussis that is cleaved to yield a 73-kDa N-terminal α-domain and a 30-kDa C-terminal β-domain. We have previously shown that a recombinant form of the β-domain of BrkA is capable of forming channels in artificial membranes. Here, we define two additional secretion determinants of BrkA. N-terminal sequencing of the 73-kDa BrkA passenger from B. pertussis and Escherichia coli revealed that BrkA has a 42-amino-acid signal peptide. In addition, deletion analysis of BrkA identified a 31- to 39-amino-acid region found immediately upstream of the β-domain that was essential for surface expression. This 31- to 39-amino-acid linker region, together with the β-domain, defines the minimal BrkA translocation unit. The linker region may also serve to anchor the BrkA passenger to the bacterial surface.

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Suzuki, Koichiro, Naoaki Shinzawa, Keisuke Ishigaki, Keiji Nakamura, Hiroyuki Abe, Aya Fukui-Miyazaki, Kazuyoshi Ikuta, and Yasuhiko Horiguchi. "Protective effects ofin vivo-expressed autotransporters againstBordetella pertussisinfection." Microbiology and Immunology 61, no. 9 (September 2017): 371–79. http://dx.doi.org/10.1111/1348-0421.12504.

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Dutta, Pinaki R., Renato Cappello, Fernando Navarro-García, and James P. Nataro. "Functional Comparison of Serine Protease Autotransporters of Enterobacteriaceae." Infection and Immunity 70, no. 12 (December 2002): 7105–13. http://dx.doi.org/10.1128/iai.70.12.7105-7113.2002.

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ABSTRACT The plasmid-encoded toxin (Pet) of enteroaggregative Escherichia coli (EAEC) belongs to a family of high-molecular-weight serine protease autotransporters of Enterobacteriaceae (SPATEs) which also includes Pic from EAEC and Shigella flexneri, EspC from enteropathogenic E. coli, EspP from enterohemorrhagic E. coli, Sat from uropathogenic E. coli, Tsh from avian pathogenic E. coli, and SepA from S. flexneri. Phylogenetic analysis shows the SPATE proteins to represent a distinct subfamily of autotransporters with amino acid identities ranging from 35 to 55%, providing a powerful resource to direct structure-function studies. In this study, we show that these related proteins are proteases with divergent substrate specificities, suggesting different functions. The cleavage profile of oligopeptides was found to be unique for each SPATE protein. The SPATEs showed proteolytic activity for several substrates, namely mucin, pepsin, human coagulation factor V, and erythroid spectrin. The cleavage of spectrin has been hypothesized as the mechanism through which Pet induces cytopathic effects. However, whereas Pet, Sat, and EspC cleaved spectrin, only Pet and Sat elicited cytopathic effects; the remaining SPATEs did not cause any morphological changes to HEp-2 cell monolayers. EspC and Pet exhibited acid-dissociable binding to HEp-2 cells. However, Pet was more efficient at entering HEp-2 cells, suggesting a basis for the different abilities of these two proteases to damage cells. Our data suggest that, despite the homologies observed among these proteins, the SPATEs have different pathogenetic functions only partly dependent on their substrate specificities.

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Kim, David S. H., Yi Chao, and Milton H. Saier. "Protein-Translocating Trimeric Autotransporters of Gram-Negative Bacteria." Journal of Bacteriology 188, no. 16 (August 15, 2006): 5655–67. http://dx.doi.org/10.1128/jb.01596-05.

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Yen, Yihfen T., Maria Kostakioti, Ian R. Henderson, and Christos Stathopoulos. "Common themes and variations in serine protease autotransporters." Trends in Microbiology 16, no. 8 (August 2008): 370–79. http://dx.doi.org/10.1016/j.tim.2008.05.003.

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Klemm, Per, Rebecca Munk Vejborg, and Orla Sherlock. "Self-associating autotransporters, SAATs: Functional and structural similarities." International Journal of Medical Microbiology 296, no. 4-5 (August 2006): 187–95. http://dx.doi.org/10.1016/j.ijmm.2005.10.002.

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Wells, Timothy J., Makrina Totsika, and Mark A. Schembri. "Autotransporters of Escherichia coli: a sequence-based characterization." Microbiology 156, no. 8 (August 1, 2010): 2459–69. http://dx.doi.org/10.1099/mic.0.039024-0.

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Autotransporter (AT) proteins are found in all Escherichia coli pathotypes and are often associated with virulence. In this study we took advantage of the large number of available E. coli genome sequences to perform an in-depth bioinformatic analysis of AT-encoding genes. Twenty-eight E. coli genome sequences were probed using an iterative approach, which revealed a total of 215 AT-encoding sequences that represented three major groups of distinct domain architecture: (i) serine protease AT proteins, (ii) trimeric AT adhesins and (iii) AIDA-I-type AT proteins. A number of subgroups were identified within each broad category, and most subgroups contained at least one characterized AT protein; however, seven subgroups contained no previously described proteins. The AIDA-I-type AT proteins represented the largest and most diverse group, with up to 16 subgroups identified from sequence-based comparisons. Nine of the AIDA-I-type AT protein subgroups contained at least one protein that possessed functional properties associated with aggregation and/or biofilm formation, suggesting a high degree of redundancy for this phenotype. The Ag43, YfaL/EhaC, EhaB/UpaC and UpaG subgroups were found in nearly all E. coli strains. Among the remaining subgroups, there was a tendency for AT proteins to be associated with individual E. coli pathotypes, suggesting that they contribute to tissue tropism or symptoms specific to different disease outcomes.

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Jong, Wouter SP, Ana Saurí, and Joen Luirink. "Extracellular production of recombinant proteins using bacterial autotransporters." Current Opinion in Biotechnology 21, no. 5 (October 2010): 646–52. http://dx.doi.org/10.1016/j.copbio.2010.07.009.

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Lawrenz, Matthew B., Jonathan D. Lenz, and Virginia L. Miller. "A Novel Autotransporter Adhesin Is Required for Efficient Colonization during Bubonic Plague." Infection and Immunity 77, no. 1 (October 20, 2008): 317–26. http://dx.doi.org/10.1128/iai.01206-08.

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ABSTRACT Many proteins secreted by the type V secretion system (autotransporters) have been linked to virulence in gram-negative bacteria. Several putative conventional autotransporters are present in the Yersinia pestis genome, but only one, YapE, is conserved in the other pathogenic Yersinia species. Here, we introduce YapE and demonstrate that it is secreted via a type V mechanism. Inactivation of yapE in Y. pestis results in decreased efficiency in colonization of tissues during bubonic infection. Coinfection with wild-type bacteria only partially compensates for this defect. Analysis of the host immune response suggests that YapE is required for either efficient colonization at the inoculation site or dissemination to draining lymph nodes. YapE also demonstrates adhesive properties capable of mediating interactions with bacteria and eukaryotic cells. These findings support a role for YapE in modulating host-pathogen interactions that are important for colonization of the mammalian host.

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Journal articles on the topic 'Autotransporters'

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