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At least 865 records · Page 48Linked to original sources
PubMed2003

Role for recombinant gamma-glutamyltransferase from Treponema denticola in glutathione metabolism.

Volatile sulfur compounds, including hydrogen sulfide (H(2)S), have been implicated in the development of periodontal disease. Glutathione is an important thiol source for H(2)S production in periodontal pockets. Our recent studies have delineated a pathway of glutathione metabolism in Treponema denticola that releases H(2)S. In this pathway, gamma-glutamyltransferase (GGT) has been proposed to catalyze the first step of glutathione degradation. We have cloned the gene of GGT from T. denticola, which contains an open reading frame of 726 bp encoding a protein of 241 amino acids. Transformation of this gene into Escherichia coli led to the expression of a recombinant protein. After purification by chromatography, the recombinant protein showed enzymatic activity typical of GGT, catalyzing the degradation of Na-gamma-glutamyl-4-nitroaniline (GNA) and the hydrolysis of glutathione, releasing glutamic acid or glutamine and cysteinylglycine. L-Cysteine is not a substrate of GGT. Importantly, GNA, when added to T. denticola, was able to compete with glutathione and inhibit the production of H(2)S, ammonia, and pyruvate. This was accompanied by the suppression of hemoxidative and hemolytic activities of the bacteria. Purified GGT was inactivated by TLCK (Nalpha-p-tosyl-L-lysine chloromethyl ketone) and proteinase K treatment. However, higher enzymatic activity was demonstrated in the presence of 2-mercaptoethanol and dithiothreitol. Our further experiments showed that the addition of recombinant GGT to Porphyromonas gingivalis, a bacterium without significant glutathione-metabolizing capacity, drastically increased the utilization of glutathione by the bacterium, producing H(2)S, ammonia, and pyruvate. This was again accompanied by enhanced bacterial hemoxidative and hemolytic activities. Together, the results suggest an important role for GGT in glutathione metabolism in oral bacteria.

Amino Acid Sequence↗
PubMed2003

Protection against syphilis correlates with specificity of antibodies to the variable regions of Treponema pallidum repeat protein K.

Syphilis has been recognized as a disease since the late 1400s, yet there is no practical vaccine available. One impediment to the development of a vaccine is the lack of understanding of multiple reinfections in humans despite the development of robust immune responses during the first episode. It has been shown that the Treponema pallidum repeat protein K (TprK) differs in seven discrete variable (V) regions in isolates and that the antibody response during infection is directed to these V regions. Immunization with TprK confers significant protection against infection with the homologous strain. We hypothesize that the antigenic diversity of TprK is involved in immune evasion, which contributes to the lack of heterologous protection. Here, using the rabbit model, we show a correlation between limited heterologous protection and tprK diversity in the challenge inoculum. We demonstrate that antibody responses to the V regions of one TprK molecule show limited cross-reactivity with heterologous TprK V regions.

Amino Acid Sequence↗
PubMed2003

Identification of a Treponema pallidum laminin-binding protein.

Host extracellular matrix (ECM) components represent ideal microbial adhesion targets that many pathogens use for colonization of tissues and initiation of infection. This study investigated the interaction of the spirochete Treponema pallidum with the ECM component laminin. To identify candidate laminin-binding adhesins, the T. pallidum genome was analyzed to predict open reading frames that encode putative outer membrane proteins, as these proteins interact directly with host ECM components. Subsequent recombinant expression of these proteins and analysis of their laminin-binding potential identified one protein, Tp0751, that demonstrated specific attachment to laminin. Tp0751 attached to laminin in a dose-dependent, saturable manner but did not attach to the ECM component collagen type I or IV or to the negative control proteins fetuin or bovine serum albumin. Sodium metaperiodate treatment of laminin reduced the Tp0751-laminin interaction in a concentration-dependent manner, suggesting that oligosaccharides play a role in this interaction. In addition, Tp0751-specific antibodies were detected in serum samples collected from both experimental and natural syphilis infections, indicating that Tp0751 is expressed in vivo during the course of infection. Collectively, these experiments identified Tp0751 as a laminin-binding protein that is expressed during infection and may be involved in attachment of T. pallidum to host tissues.

Adhesins, Bacterial↗
PubMed2004

Induction of de novo subcortical actin filament assembly by Treponema denticola major outer sheath protein.

Treponema denticola and its major outer sheath protein (Msp) induce actin reorganization in fibroblasts. We adapted a barbed-end labeling/imaging assay to monitor Msp-induced subcortical actin filament assembly in neutrophils and fibroblasts. Msp, at an actin-reorganizing concentration, inhibited migration of these dissimilar cell types, whose cytoskeletal functions in locomotion and phagocytosis are crucial for immunity and healing of peripheral infections.

Actin Cytoskeleton↗
PubMed2004

Multiple functions of the leucine-rich repeat protein LrrA of Treponema denticola.

The gene lrrA, encoding a leucine-rich repeat protein, LrrA, that contains eight consensus tandem repeats of 23 amino acid residues, has been identified in Treponema denticola ATCC 35405. A leucine-rich repeat is a generally useful protein-binding motif, and proteins containing this repeat are typically involved in protein-protein interactions. Southern blot analysis demonstrated that T. denticola ATCC 35405 expresses the lrrA gene, but the gene was not identified in T. denticola ATCC 33520. In order to analyze the functions of LrrA in T. denticola, an lrrA-inactivated mutant of strain ATCC 35405 and an lrrA gene expression transformant of strain ATCC 33520 were constructed. Characterization of the mutant and transformant demonstrated that LrrA is associated with the extracytoplasmic fraction of T. denticola and expresses multifunctional properties. It was demonstrated that the attachment of strain ATCC 35405 to HEp-2 cell cultures and coaggregation with Tannerella forsythensis were attenuated by the lrrA mutation. In addition, an in vitro binding assay demonstrated specific binding of LrrA to a portion of the Tannerella forsythensis leucine-rich repeat protein, BspA, which is mediated by the N-terminal region of LrrA. It was also observed that the lrrA mutation caused a reduction of swarming in T. denticola ATCC 35405 and consequently attenuated tissue penetration. These results suggest that the leucine-rich repeat protein LrrA plays a role in the attachment and penetration of human epithelial cells and coaggregation with Tannerella forsythensis. These properties may play important roles in the virulence of T. denticola.

Amino Acid Sequence↗
PubMed2005

Demonstration of factor H-like protein 1 binding to Treponema denticola, a pathogen associated with periodontal disease in humans.

Treponema denticola is an important contributor to periodontal disease. In this study we investigated the ability of T. denticola to bind the complement regulatory proteins factor H and factor H-like protein 1 (FHL-1). The binding of these proteins has been demonstrated to facilitate evasion of the alternative complement cascade and/or to play a role in adherence and invasion. Here we demonstrate that T. denticola specifically binds FHL-1 via a 14-kDa, surface-exposed protein that we designated FhbB. Consistent with its FHL-1 binding specificity, FhbB binds only to factor H recombinant fragments spanning short consensus repeats (SCRs) 1 to 7 (H7 construct) and not to SCR constructs spanning SCRs 8 to 15 and 16 to 20. Binding of H7 to FhbB was inhibited by heparin. The specific involvement of SCR 7 in the interaction was demonstrated using an H7 mutant (H7AB) in which specific charged residues in SCR 7 were replaced by alanine. This construct lost FhbB binding ability. Analyses of the ability of FHL-1 bound to the surface of T. denticola to serve as a cofactor for factor I-mediated cleavage of C3b revealed that C3b is cleaved in an FHL-1/factor I-independent manner, perhaps by an unidentified protease. Based on the data presented here, we hypothesize that the primary function of FHL-1 binding by T. denticola might be to facilitate adherence to FHL-1 present on anchorage-dependent cells and in the extracellular matrix.

Bacterial Proteins↗
PubMed2005

Defining the interaction of the Treponema pallidum adhesin Tp0751 with laminin.

Various invasive pathogens attach to host tissues via the extracellular matrix component laminin, the major glycoprotein found within basement membranes. Previous investigations identified the laminin-binding adhesin Tp0751 within the spirochete bacterium Treponema pallidum. In the current study, Tp0751 was shown to attach to a variety of laminin isoforms that are widely distributed throughout the host, including laminins 1, 2, 4, 8, and 10. Such universal attachment is conducive for an adhesin present within a highly invasive pathogen that encounters a variety of tissue sites during the course of infection. Additional studies systematically identified the amino acid residues within Tp0751 that contribute to laminin binding using synthetic peptides designed from the mature protein sequence. The minimum laminin-binding region of the adhesin was localized to 10 amino acids; peptides containing these residues inhibited attachment of Tp0751 and T. pallidum to laminin. Further, Tp0751-specific antibodies inhibited attachment of T. pallidum to laminin. This study furthers our knowledge of the interaction of T. pallidum with laminin, an association that is proposed to facilitate bacterial traversal of basement membranes and subsequent entry into the circulation and tissue invasion. As such, these investigations will reveal new targets for possible prevention of bacterial dissemination and establishment of chronic infection.

Adhesins, Bacterial↗
PubMed2005

Mutagenesis of a novel gene in the prcA-prtP protease locus affects expression of Treponema denticola membrane complexes.

A novel gene was identified in the Treponema denticola prcA-prtP protease operon. Strains with mutations in either the prcA-prtP or the msp region showed altered expression of a product(s) of the other locus. Together, these results provide information on the assembly of outer membrane complexes involved in T. denticola interaction with host cells and tissue.

Bacterial Proteins↗
PubMed2005

A monoclonal antibody that conveys in vitro killing and partial protection in experimental syphilis binds a phosphorylcholine surface epitope of Treponema pallidum.

Immunization with purified Treponema pallidum outer membrane vesicles (OMV) has previously resulted in high-titer complement-dependent serum bactericidal activity. In this study, OMV immunization resulted in the isolation of a monoclonal antibody, M131, with complement-dependent killing activity. Passive immunization of rabbits with M131 administered intravenously conferred significant immunity demonstrated by the failure of syphilitic lesions to appear at 29% of intradermal challenge sites (7/24) and a mean delay of approximately 8 days to lesion appearance at the remaining sites (17/24). M131 not only bound to OMV and to the surfaces of intact motile T. pallidum cells but also bound to organisms whose outer membranes were removed, indicating both surface and subsurface locations for the killing target. This target was determined to be a T. pallidum lipid. Lipid extracted from T. pallidum and made into liposomes bound M131. Reverse-phase high-pressure liquid chromatography separation and fraction collection mass spectrometry (LC-MS+) of T. pallidum lipid showed that the target of M131 was phosphorylcholine. M131 binding required both liposome formation and a critical concentration of phospholipid containing phosphorylcholine, suggesting that the epitope has both a conformational and a compositional requirement. M131 did not react with red blood cells, which have phosphorylcholine-containing lipids in their exterior membrane leaflets, or with Venereal Disease Research Laboratory antigen that also contains phosphorylcholine, further indicating the specificity of M131. This is the first physical demonstration of an antigen on the T. pallidum surface and indication that such a surface antigen can be a target of immunity.

Animals↗
PubMed2005

Immunological evaluation and cellular location analysis of the TprI antigen of Treponema pallidum subsp. pallidum.

The TprI antigen of Treponema pallidum subsp. pallidum is a putative virulence factor predicted to be located in the outer membrane of the syphilis spirochete. In this study, we analyzed the immune response against TprI and its subunits in sera collected both from rabbits experimentally infected with the Nichols strain and from patients with syphilis, showing a different pattern of reactivity toward the antigen in these two groups of samples. The protective ability of recombinant TprI and its hypothetical outer membrane location were also investigated. Although no rabbit was protected after challenge, immunoelectron microscopy results, to be further investigated, were compatible with the outer membrane location of the antigen.

Animals↗
PubMed2005

Genome scale identification of Treponema pallidum antigens.

Antibody responses for 882 of the 1,039 proteins in the proteome of Treponema pallidum were examined. Sera collected from infected rabbits were used to systematically identify 106 antigenic proteins, including 22 previously identified antigens and 84 novel antigens. Additionally, sera collected from rabbits throughout the course of infection demonstrated a progression in the breadth and intensity of humoral immunoreactivity against a representative panel of T. pallidum antigens.

Animals↗
PubMed2006

TprK sequence diversity accumulates during infection of rabbits with Treponema pallidum subsp. pallidum Nichols strain.

The tprK gene in Treponema pallidum undergoes antigenic variation. In all T. pallidum isolates examined to date, except the Nichols type strain, heterogeneous tprK sequences have been identified. This heterogeneity is localized to seven variable (V) regions, and tprK sequence diversity accumulates with serial passage in naïve rabbits. The T. pallidum Nichols genome described a single tprK sequence, and after decades of independent passage, only minor tprK sequence diversity is seen among the Nichols strains from different laboratories. We hypothesized that T. pallidum Nichols is capable of only limited tprK diversification. To address this hypothesis, we passaged the T. pallidum Nichols strain in naïve rabbits at the peak of infection (rapid passage) or after the adaptive immune response had cleared most organisms in vivo (slow passage). After 22 rapid passages (9- to 10-day intervals), no tprK V region sequence changes were observed. In contrast, after two slow passages (30- to 35-day intervals), three V regions had sequences that were completely different from that of the original inoculum. New sequences were observed in all seven V regions by the fifth slow passage. In contrast to the rapid-passaged Nichols strain, rapid-passaged Chicago C, a clonal strain isolated from the highly diverse parent Chicago strain, developed significant tprK diversification. These findings suggest that tprK variation can occur, but at a lower rate, in Nichols and that immune pressure may be required for accumulation of bacteria with diverse tprK sequences. Adaptation to growth in rabbits may explain the limited repertoire of V region sequences seen in the Nichols strain.

Amino Acid Sequence↗
PubMed2006

Modulation of human neutrophil functions in vitro by Treponema denticola major outer sheath protein.

In this study of human polymorphonuclear leukocytes (PMNs), pretreatment with Treponema denticola major outer sheath protein (Msp) inhibited formyl-methionyl-leucyl-phenylalanine (fMLP)-induced chemotaxis, phagocytosis of immunoglobulin G-coated microspheres, fMLP-stimulated calcium transients, and actin assembly. Msp neither altered oxidative responses to phorbol myristate or fMLP nor induced apoptosis. Msp selectively impairs chemotaxis and phagocytosis by impacting the PMN cytoskeleton.

Bacterial Proteins↗
PubMed1974

Localization of spirochetes with the structural characteristics of Treponema hyodysenteriae in the lesions of swine dysentery.

The large intestines of pigs with swine dysentery were examined by phase, light, and electron microscopy at intervals up to 11 days after oral inoculation with mucosal scrapings from infected pigs. Large spirochetes with the structural characteristics of Treponema hyodysenteriae were found only in infected pigs and were first observed in small numbers in the lumen of the large intestine 2 days after inoculation. Numerous large spirochetes were present on the luminal surface and in mucosal crypts as lesions developed. Degenerative changes were first observed in the apical portion of epithelial cells in close contact with large spirochetes. These large spirochetes were found intact in goblet cells and epithelial cells in the early stages of the disease and were numerous within degenerating epithelial cells as lesions became more advanced. Invasion beyond the lamina propria was not detected. These observations demonstrated the relationship between pathogenic large spirochetes and the mucosa of the large intestine in a specific disease, swine dysentery.

Animals↗
PubMed1974

In vitro lymphocyte response to Treponema refringens im human syphilis.

The response of lymphocytes from patients with syphilis and normal subjects was studied in vitro by using phytohemagglutinin (PHA), pokeweed mitogen (PWM), streptolysin O (SLO), and a preparation of Treponema refringens. Normal lymphocytes exhibited a dose-response curve to treponemes. Although lymphocytes from patients with primary and secondary syphilis responded normally to PHA and PWM, their response to SLO was suppressed and they failed to show significant stimulation by treponemes. Serum from syphilitic patients did not affect normal lymphocytes, and culturing lymphocytes from patients with syphilis in normal serum did not restore their responsiveness. Six to 10 weeks after syphilitic patients had been treated, the degree of stimulation by treponemes was the same as for normal subjects. These data give indirect support to the hypothesis that immunological suppression occurs during active infection with T. pallidum.

Antigens, Bacterial↗
PubMed1974

Detection of nonspecific resistance to Listeria monocytogenes in rabbits infected with Treponema pallidum.

Several lines of evidence suggest that cell-mediated immunity (CMI) is suppressed in the early stages of infection caused by Treponema pallidum and becomes activated at the time that latency is induced. In the studies reported in this paper, rabbits were infected intravenously with T. pallidum and subsequently challenged with Listeria monocytogenes. Enhanced ability to suppress the growth of Listeria was detected in their livers between 3 and 5 weeks after infection with T. pallidum, corresponding to the onset and regression of the generalized syphilitic eruption. A second infection of T. pallidum 4 weeks after the first, at a time when suppression was beginning to wane, prolonged the listericidal activity. These observations support the hypothesis that infection by T. pallidum stimulates CMI, which, in turn, may play a role in inducing latency.

Animals↗