Orthologous clusters and OMP prediction heat map. Orthologous clusters from each borrelial genome... more Orthologous clusters and OMP prediction heat map. Orthologous clusters from each borrelial genome organized in unique columns as a heat map. Green blocks in each column indicate Borrelia proteins predicted to be OMPs, while Borrelia proteins not passing all OMP prediction filters are represented by red blocks. Black blocks indicate an orthologous protein was not identified in a given borrelial genome. These data were then subjected to a two-way hierarchical clustering examining orthologs (column blocks) by genomes (row blocks). Unique cluster IDs are shown at the bottom of the figure and correspond to the cluster numbers listed in Additional file 5: Table S4. Genomes analyzed are listed at right. (PDF 18 kb)
Predictions for all ORFs. Lipoprotein, trans-membrane ι-helix, cellular localization, and β-barre... more Predictions for all ORFs. Lipoprotein, trans-membrane ι-helix, cellular localization, and β-barrel prediction data collected for all ORFs for all genomes analyzed. (XLS 5022 kb)
Orthologous clusters. List of predicted orthologous clusters utilized to generate heat maps. (XLS... more Orthologous clusters. List of predicted orthologous clusters utilized to generate heat maps. (XLSX 667 kb)
In recent years, considerable progress has been made in topologically and functionally characteri... more In recent years, considerable progress has been made in topologically and functionally characterizing integral outer membrane proteins (OMPs) of subspecies , the syphilis spirochete, and identifying its surface-exposed β-barrel domains. Extracellular loops in OMPs of Gram-negative bacteria are known to be highly variable. We examined the sequence diversity of β-barrel-encoding regions of , , and in 31 specimens from Cali, Colombia; San Francisco, California; and the Czech Republic and compared them to allelic variants in the 41 reference genomes in the NCBI database. To establish a phylogenetic framework, we used 0548 () genotyping and sequences to assign strains to the Nichols or SS14 clades. We found that (i) β-barrels in clinical strains could be grouped according to allelic variants in subsp. reference genomes; (ii) for all three OMP loci, clinical strains within the Nichols or SS14 clades often harbored β-barrel variants that differed from the Nichols and SS14 reference strains...
The major outer sheath protein (MOSP) is a prominent constituent of the cell envelope of Treponem... more The major outer sheath protein (MOSP) is a prominent constituent of the cell envelope of Treponema denticola (TDE) and one of its principal virulence determinants. Bioinformatics predicts that MOSP consists of N- and C-terminal domains, MOSP(N) and MOSP(C). Biophysical analysis of constructs refolded in vitro demonstrated that MOSP(C), previously shown to possess porin activity, forms amphiphilic trimers, while MOSP(N) forms an extended hydrophilic monomer. In TDE and E. coli expressing MOSP with a PelB signal sequence (PelB-MOSP), MOSP(C) is OM-embedded and surface-exposed, while MOSP(N) resides in the periplasm. Immunofluorescence assay, surface proteolysis, and novel cell fractionation schemes revealed that MOSP in TDE exists as outer membrane (OM) and periplasmic trimeric conformers; PelB-MOSP, in contrast, formed only OM-MOSP trimers. Although both conformers form hetero-oligomeric complexes in TDE, only OM-MOSP associates with dentilisin. Mass spectrometry (MS) indicated that ...
Jarisch-Herzheimer reaction was positively associated with treatment regimen prognosis (p<0.05). ... more Jarisch-Herzheimer reaction was positively associated with treatment regimen prognosis (p<0.05). Conclusion Our serological data demonstrate that this ceftriaxone regimen is more effective than the currently recommended benzathine penicillin regimen for early syphilis in non-pregnant, immunocompetent patients, especially for secondary syphilis.
The past two decades have seen a worldwide resurgence in infections caused by Treponema pallidum ... more The past two decades have seen a worldwide resurgence in infections caused by Treponema pallidum subsp. pallidum, the syphilis spirochete. The well-recognized capacity of the syphilis spirochete for early dissemination and immune evasion has earned it the designation 'the stealth pathogen'. Despite the many hurdles to studying syphilis pathogenesis, most notably the inability to culture and to genetically manipulate T. pallidum, in recent years, considerable progress has been made in elucidating the structural, physiological, and regulatory facets of T. pallidum pathogenicity. In this Review, we integrate this eclectic body of information to garner fresh insights into the highly successful parasitic lifestyles of the syphilis spirochete and related pathogenic treponemes.
Background: Similar to Gram-negative organisms, Borrelia spirochetes are dual-membrane organisms ... more Background: Similar to Gram-negative organisms, Borrelia spirochetes are dual-membrane organisms with both an inner and outer membrane. Although the outer membrane contains integral membrane proteins, few of the borrelial outer membrane proteins (OMPs) have been identified and characterized to date. Therefore, we utilized a consensus computational network analysis to identify novel borrelial OMPs. Results: Using a series of computer-based algorithms, we selected all protein-encoding sequences predicted to be OM-localized and/or to form β-barrels in the borrelial OM. Using this system, we identified 41 potential OMPs from B. burgdorferi and characterized three (BB0838, BB0405, and BB0406) to confirm that our computer-based methodology did, in fact, identify borrelial OMPs. Triton X-114 phase partitioning revealed that BB0838 is found in the detergent phase, which would be expected of a membrane protein. Proteolysis assays indicate that BB0838 is partially sensitive to both proteinase K and trypsin, further indicating that BB0838 is surface-exposed. Consistent with a prior study, we also confirmed that BB0405 is surface-exposed and associates with the borrelial OM. Furthermore, we have shown that BB0406, the product of a co-transcribed downstream gene, also encodes a novel, previously uncharacterized borrelial OMP. Interestingly, while BB0406 has several physicochemical properties consistent with it being an OMP, it was found to be resistant to surface proteolysis. Consistent with BB0405 and BB0406 being OMPs, both were found to be capable of incorporating into liposomes and exhibit pore-forming activity, suggesting that both proteins are porins. Lastly, we expanded our computational analysis to identify OMPs from other borrelial organisms, including both Lyme disease and relapsing fever spirochetes. Conclusions: Using a consensus computer algorithm, we generated a list of candidate OMPs for both Lyme disease and relapsing fever spirochetes and determined that three of the predicted B. burgdorferi proteins identified were indeed novel borrelial OMPs. The combined studies have identified putative spirochetal OMPs that can now be examined for their roles in virulence, physiology, and disease pathogenesis. Importantly, the studies described in this report provide a framework by which OMPs from any human pathogen with a diderm ultrastructure could be cataloged to identify novel virulence factors and vaccine candidates.
The noncultivable spirochete Treponema pallidum subspecies pallidum (T. pallidum) is the etiologi... more The noncultivable spirochete Treponema pallidum subspecies pallidum (T. pallidum) is the etiological agent of venereal syphilis. In contrast to the outer membranes (OMs) of gram-negative bacteria, the OM of T. pallidum lacks lipopolysaccharide, contains a paucity of integral membrane proteins, and is extremely labile. The lability of the T. pallidum OM greatly hinders efforts to localize the bacterium&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s rare outer membrane proteins (OMPs). To circumvent this problem, we developed the gel microdroplet method in which treponemes are encapsulated in porous agarose beads and then probed with specific antibodies in the absence or presence of low concentrations of the non-ionic detergent Triton X-100. To demonstrate the general utility of this method for surface localization of any T. pallidum antigen, herein we describe a protocol for immunolabeling of encapsulated treponemes using antibodies directed against the β-barrel and POTRA domains of TP0326, the spirochete&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s BamA ortholog.
Figure 4C should appear as shown below. An image containing the two rightmost lanes from Fig. 4B ... more Figure 4C should appear as shown below. An image containing the two rightmost lanes from Fig. 4B was inadvertently used to generate the top left panel in Fig. 4C (Intact T. denticola Ϫ/ϩ PK treatment immunoblotted with anti-Msp). In addition, the middle and bottom right panels (Disrupted T. denticola Ϫ/ϩ PK treatment immunoblotted with anti-TroA and anti-FlaA, respectively) were inadvertently switched in the composite figure. The corrected figure shows that, as stated in Results, Msp is largely inaccessible to proteinase K because it is not on the surface of the treponeme as opposed to being protease resistant.
The legend to Fig. 6E should read as follows. "Immunoblot of unboiled, recombinant TprC N and Tpr... more The legend to Fig. 6E should read as follows. "Immunoblot of unboiled, recombinant TprC N and TprC C separated by SDS-PAGE and then probed with rabbit anti-TprC Fl polyclonal antiserum (asterisk indicates TprC C trimer)." Page 2331: The legend to Fig. 7C should read as follows. "Reactivities of the anti-TprC Fl antiserum and IRS used for the opsonophagocytosis assays in Fig. 5 against TprC N and folded, unboiled TprC C. The immunoblot shown in Fig. 6E was stripped and reprobed with IRS. Bottom and top panels show the reactivity of TprC N and TprC C monomers with anti-TprC Fl antiserum (reproduced from Fig. 6E) and IRS, respectively."
We recently demonstrated that TP0326 is a bona fide rare outer membrane protein (OMP) in Treponem... more We recently demonstrated that TP0326 is a bona fide rare outer membrane protein (OMP) in Treponema pallidum (Tp) and that it possesses characteristic BamA bipartite topology. Herein we used immunofluorescence analysis (IFA) to show that only the β-barrel domain of TP0326 contains surface-exposed epitopes in intact Tp. Using the solved structure of Neisseria gonorrhoeae BamA, we generated a homology model of full-length TP0326. Although the model predicts a typical BamA fold, the β-barrel harbors features not described in other BamAs. Structural modeling predicted that a dome comprised of three large extracellular loops, L4, L6 and L7, covers the barrel's extracellular opening. L4, the dome's major surface-accessible loop, contains mainly charged residues, while L7 is largely neutral and contains a polyserine tract in a two-tiered conformation. L6 projects into the β-barrel but lacks the VRGF/Y motif that anchors L6 within other BamA. IFA and opsonophagocytosis assay revealed...
Background: TP0453 is the only lipoprotein in T. pallidum known to be associated with the outer m... more Background: TP0453 is the only lipoprotein in T. pallidum known to be associated with the outer membrane (OM). Results: TP0453 adopts closed and open conformations; opening is associated with membrane sensing/insertion. Conclusion: Two amphipathic helices are responsible for amphiphilic behavior of the molecule. Significance: TP0453 is a novel bacterial lipoprotein whose amphiphilic behavior may contribute to T. pallidum OM biogenesis.
The Borrelia burgdorferi outer membrane (OM) contains numerous surface-exposed lipoproteins but a... more The Borrelia burgdorferi outer membrane (OM) contains numerous surface-exposed lipoproteins but a relatively low density of integral OM proteins (OMPs). Few membrane-spanning OMPs of B. burgdorferi have been definitively identified, and none are well characterized structurally. Here, we provide evidence that the borrelial OMP P66, a known adhesin with pore-forming activity, forms a β-barrel in the B. burgdorferi OM. Multiple computer-based algorithms predict that P66 forms a β-barrel with either 22 or 24 transmembrane domains. According to our predicted P66 topology, a lysine residue (K487) known to be sensitive to trypsin cleavage is located within a surface-exposed loop. When we aligned the mature P66 amino acid sequences from B. burgdorferi and B. garinii , we found that K487 was present only in the B. burgdorferi P66 protein sequence. When intact cells from each strain were treated with trypsin, only B. burgdorferi P66 was trypsin sensitive, indicating that K487 is surface expos...
In urban India, now a day's large number of multi story RC framed residential buildings are const... more In urban India, now a day's large number of multi story RC framed residential buildings are constructing with different ideas like soft storey or without soft story. The present paper investigates about the performance of non linear analysis of different RC framed models such as bare frame and open first storey and without soft storey. A soft story is an apartment building of three or more story located over a ground level with large openings such as parking area and soft storey is also called as open first storey. The study is carried out on G+4 storey buildings were considered as residential building which is located on zone III and the push over analysis and time history analysis are performed in the present study using sap 2000 v15.1.0 software. User defined hinge are assigned for beams and columns. Seismic performances of the frames were compared and effects of using different models were investigated. Results show a general changing pattern in performance point, capacity curve, demand spectrum curve.
We previously identified T. pallidum repeat proteins TprC/D, TprF, and TprI as candidate outer me... more We previously identified T. pallidum repeat proteins TprC/D, TprF, and TprI as candidate outer membrane proteins (OMPs) and subsequently demonstrated that TprC is not only a rare OMP but also forms trimers and has porin activity. We also reported that TprC contains N- and C-terminal domains (TprCN and TprCC) orthologous to regions in the major outer sheath protein (MOSPN and MOSPC) of T. denticola and that TprCC is solely responsible for β-barrel formation, trimerization, and porin function by the full-length protein. Herein, we show that TprI also possesses bipartite architecture, trimeric structure, and porin function and that the MOSPC-like domains of native TprC and TprI are surface-exposed in T. pallidum while their MOSPN-like domains are tethered within the periplasm. TprF, which does not contain a MOSPC-like domain, lacks amphiphilicity and porin activity, adopts an extended inflexible structure, and in T. pallidum, is tightly bound to the protoplasmic cylinder. By thermal de...
Definitive identification of Treponema pallidum (Tp) rare outer membrane proteins (OMPs) has long... more Definitive identification of Treponema pallidum (Tp) rare outer membrane proteins (OMPs) has long eluded researchers. TP0326, the sole protein in Tp with sequence homology to a Gramnegative OMP, belongs to the BamA family of proteins essential for OM biogenesis. Structural modeling predicted that five polypeptide transport-associated (POTRA) domains comprise the Nterminus of TP0326, while the C-terminus forms an 18-stranded amphipathic β-barrel. Circular dichroism, heat-modifiability by SDS-PAGE, Triton X-114 phase partitioning and liposome incorporation supported these topological predictions and confirmed that the β-barrel is responsible for the native protein's amphiphilicity. Expression analyses revealed that native TP0326 is expressed at low abundance, while a protease-surface accessibility assay confirmed surface exposure. Size-exclusion chromatography and blue native polyacrylamide gel electrophoresis revealed a modular Bam complex in Tp considerably larger than that of E. coli. Non-orthologous ancillary factors and self-association of TP0326 via its β-barrel may both contribute to the Bam complex. Tp-infected rabbits mount a vigorous antibody response to both POTRA and β-barrel portions of TP0326, whereas humans with secondary syphilis respond predominantly to POTRA. The syphilis spirochete appears to have devised a stratagem for harnessing the Bam pathway while satisfying its need to limit surface antigenicity.
Orthologous clusters and OMP prediction heat map. Orthologous clusters from each borrelial genome... more Orthologous clusters and OMP prediction heat map. Orthologous clusters from each borrelial genome organized in unique columns as a heat map. Green blocks in each column indicate Borrelia proteins predicted to be OMPs, while Borrelia proteins not passing all OMP prediction filters are represented by red blocks. Black blocks indicate an orthologous protein was not identified in a given borrelial genome. These data were then subjected to a two-way hierarchical clustering examining orthologs (column blocks) by genomes (row blocks). Unique cluster IDs are shown at the bottom of the figure and correspond to the cluster numbers listed in Additional file 5: Table S4. Genomes analyzed are listed at right. (PDF 18 kb)
Predictions for all ORFs. Lipoprotein, trans-membrane ι-helix, cellular localization, and β-barre... more Predictions for all ORFs. Lipoprotein, trans-membrane ι-helix, cellular localization, and β-barrel prediction data collected for all ORFs for all genomes analyzed. (XLS 5022 kb)
Orthologous clusters. List of predicted orthologous clusters utilized to generate heat maps. (XLS... more Orthologous clusters. List of predicted orthologous clusters utilized to generate heat maps. (XLSX 667 kb)
In recent years, considerable progress has been made in topologically and functionally characteri... more In recent years, considerable progress has been made in topologically and functionally characterizing integral outer membrane proteins (OMPs) of subspecies , the syphilis spirochete, and identifying its surface-exposed β-barrel domains. Extracellular loops in OMPs of Gram-negative bacteria are known to be highly variable. We examined the sequence diversity of β-barrel-encoding regions of , , and in 31 specimens from Cali, Colombia; San Francisco, California; and the Czech Republic and compared them to allelic variants in the 41 reference genomes in the NCBI database. To establish a phylogenetic framework, we used 0548 () genotyping and sequences to assign strains to the Nichols or SS14 clades. We found that (i) β-barrels in clinical strains could be grouped according to allelic variants in subsp. reference genomes; (ii) for all three OMP loci, clinical strains within the Nichols or SS14 clades often harbored β-barrel variants that differed from the Nichols and SS14 reference strains...
The major outer sheath protein (MOSP) is a prominent constituent of the cell envelope of Treponem... more The major outer sheath protein (MOSP) is a prominent constituent of the cell envelope of Treponema denticola (TDE) and one of its principal virulence determinants. Bioinformatics predicts that MOSP consists of N- and C-terminal domains, MOSP(N) and MOSP(C). Biophysical analysis of constructs refolded in vitro demonstrated that MOSP(C), previously shown to possess porin activity, forms amphiphilic trimers, while MOSP(N) forms an extended hydrophilic monomer. In TDE and E. coli expressing MOSP with a PelB signal sequence (PelB-MOSP), MOSP(C) is OM-embedded and surface-exposed, while MOSP(N) resides in the periplasm. Immunofluorescence assay, surface proteolysis, and novel cell fractionation schemes revealed that MOSP in TDE exists as outer membrane (OM) and periplasmic trimeric conformers; PelB-MOSP, in contrast, formed only OM-MOSP trimers. Although both conformers form hetero-oligomeric complexes in TDE, only OM-MOSP associates with dentilisin. Mass spectrometry (MS) indicated that ...
Jarisch-Herzheimer reaction was positively associated with treatment regimen prognosis (p<0.05). ... more Jarisch-Herzheimer reaction was positively associated with treatment regimen prognosis (p<0.05). Conclusion Our serological data demonstrate that this ceftriaxone regimen is more effective than the currently recommended benzathine penicillin regimen for early syphilis in non-pregnant, immunocompetent patients, especially for secondary syphilis.
The past two decades have seen a worldwide resurgence in infections caused by Treponema pallidum ... more The past two decades have seen a worldwide resurgence in infections caused by Treponema pallidum subsp. pallidum, the syphilis spirochete. The well-recognized capacity of the syphilis spirochete for early dissemination and immune evasion has earned it the designation 'the stealth pathogen'. Despite the many hurdles to studying syphilis pathogenesis, most notably the inability to culture and to genetically manipulate T. pallidum, in recent years, considerable progress has been made in elucidating the structural, physiological, and regulatory facets of T. pallidum pathogenicity. In this Review, we integrate this eclectic body of information to garner fresh insights into the highly successful parasitic lifestyles of the syphilis spirochete and related pathogenic treponemes.
Background: Similar to Gram-negative organisms, Borrelia spirochetes are dual-membrane organisms ... more Background: Similar to Gram-negative organisms, Borrelia spirochetes are dual-membrane organisms with both an inner and outer membrane. Although the outer membrane contains integral membrane proteins, few of the borrelial outer membrane proteins (OMPs) have been identified and characterized to date. Therefore, we utilized a consensus computational network analysis to identify novel borrelial OMPs. Results: Using a series of computer-based algorithms, we selected all protein-encoding sequences predicted to be OM-localized and/or to form β-barrels in the borrelial OM. Using this system, we identified 41 potential OMPs from B. burgdorferi and characterized three (BB0838, BB0405, and BB0406) to confirm that our computer-based methodology did, in fact, identify borrelial OMPs. Triton X-114 phase partitioning revealed that BB0838 is found in the detergent phase, which would be expected of a membrane protein. Proteolysis assays indicate that BB0838 is partially sensitive to both proteinase K and trypsin, further indicating that BB0838 is surface-exposed. Consistent with a prior study, we also confirmed that BB0405 is surface-exposed and associates with the borrelial OM. Furthermore, we have shown that BB0406, the product of a co-transcribed downstream gene, also encodes a novel, previously uncharacterized borrelial OMP. Interestingly, while BB0406 has several physicochemical properties consistent with it being an OMP, it was found to be resistant to surface proteolysis. Consistent with BB0405 and BB0406 being OMPs, both were found to be capable of incorporating into liposomes and exhibit pore-forming activity, suggesting that both proteins are porins. Lastly, we expanded our computational analysis to identify OMPs from other borrelial organisms, including both Lyme disease and relapsing fever spirochetes. Conclusions: Using a consensus computer algorithm, we generated a list of candidate OMPs for both Lyme disease and relapsing fever spirochetes and determined that three of the predicted B. burgdorferi proteins identified were indeed novel borrelial OMPs. The combined studies have identified putative spirochetal OMPs that can now be examined for their roles in virulence, physiology, and disease pathogenesis. Importantly, the studies described in this report provide a framework by which OMPs from any human pathogen with a diderm ultrastructure could be cataloged to identify novel virulence factors and vaccine candidates.
The noncultivable spirochete Treponema pallidum subspecies pallidum (T. pallidum) is the etiologi... more The noncultivable spirochete Treponema pallidum subspecies pallidum (T. pallidum) is the etiological agent of venereal syphilis. In contrast to the outer membranes (OMs) of gram-negative bacteria, the OM of T. pallidum lacks lipopolysaccharide, contains a paucity of integral membrane proteins, and is extremely labile. The lability of the T. pallidum OM greatly hinders efforts to localize the bacterium&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s rare outer membrane proteins (OMPs). To circumvent this problem, we developed the gel microdroplet method in which treponemes are encapsulated in porous agarose beads and then probed with specific antibodies in the absence or presence of low concentrations of the non-ionic detergent Triton X-100. To demonstrate the general utility of this method for surface localization of any T. pallidum antigen, herein we describe a protocol for immunolabeling of encapsulated treponemes using antibodies directed against the β-barrel and POTRA domains of TP0326, the spirochete&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s BamA ortholog.
Figure 4C should appear as shown below. An image containing the two rightmost lanes from Fig. 4B ... more Figure 4C should appear as shown below. An image containing the two rightmost lanes from Fig. 4B was inadvertently used to generate the top left panel in Fig. 4C (Intact T. denticola Ϫ/ϩ PK treatment immunoblotted with anti-Msp). In addition, the middle and bottom right panels (Disrupted T. denticola Ϫ/ϩ PK treatment immunoblotted with anti-TroA and anti-FlaA, respectively) were inadvertently switched in the composite figure. The corrected figure shows that, as stated in Results, Msp is largely inaccessible to proteinase K because it is not on the surface of the treponeme as opposed to being protease resistant.
The legend to Fig. 6E should read as follows. "Immunoblot of unboiled, recombinant TprC N and Tpr... more The legend to Fig. 6E should read as follows. "Immunoblot of unboiled, recombinant TprC N and TprC C separated by SDS-PAGE and then probed with rabbit anti-TprC Fl polyclonal antiserum (asterisk indicates TprC C trimer)." Page 2331: The legend to Fig. 7C should read as follows. "Reactivities of the anti-TprC Fl antiserum and IRS used for the opsonophagocytosis assays in Fig. 5 against TprC N and folded, unboiled TprC C. The immunoblot shown in Fig. 6E was stripped and reprobed with IRS. Bottom and top panels show the reactivity of TprC N and TprC C monomers with anti-TprC Fl antiserum (reproduced from Fig. 6E) and IRS, respectively."
We recently demonstrated that TP0326 is a bona fide rare outer membrane protein (OMP) in Treponem... more We recently demonstrated that TP0326 is a bona fide rare outer membrane protein (OMP) in Treponema pallidum (Tp) and that it possesses characteristic BamA bipartite topology. Herein we used immunofluorescence analysis (IFA) to show that only the β-barrel domain of TP0326 contains surface-exposed epitopes in intact Tp. Using the solved structure of Neisseria gonorrhoeae BamA, we generated a homology model of full-length TP0326. Although the model predicts a typical BamA fold, the β-barrel harbors features not described in other BamAs. Structural modeling predicted that a dome comprised of three large extracellular loops, L4, L6 and L7, covers the barrel's extracellular opening. L4, the dome's major surface-accessible loop, contains mainly charged residues, while L7 is largely neutral and contains a polyserine tract in a two-tiered conformation. L6 projects into the β-barrel but lacks the VRGF/Y motif that anchors L6 within other BamA. IFA and opsonophagocytosis assay revealed...
Background: TP0453 is the only lipoprotein in T. pallidum known to be associated with the outer m... more Background: TP0453 is the only lipoprotein in T. pallidum known to be associated with the outer membrane (OM). Results: TP0453 adopts closed and open conformations; opening is associated with membrane sensing/insertion. Conclusion: Two amphipathic helices are responsible for amphiphilic behavior of the molecule. Significance: TP0453 is a novel bacterial lipoprotein whose amphiphilic behavior may contribute to T. pallidum OM biogenesis.
The Borrelia burgdorferi outer membrane (OM) contains numerous surface-exposed lipoproteins but a... more The Borrelia burgdorferi outer membrane (OM) contains numerous surface-exposed lipoproteins but a relatively low density of integral OM proteins (OMPs). Few membrane-spanning OMPs of B. burgdorferi have been definitively identified, and none are well characterized structurally. Here, we provide evidence that the borrelial OMP P66, a known adhesin with pore-forming activity, forms a β-barrel in the B. burgdorferi OM. Multiple computer-based algorithms predict that P66 forms a β-barrel with either 22 or 24 transmembrane domains. According to our predicted P66 topology, a lysine residue (K487) known to be sensitive to trypsin cleavage is located within a surface-exposed loop. When we aligned the mature P66 amino acid sequences from B. burgdorferi and B. garinii , we found that K487 was present only in the B. burgdorferi P66 protein sequence. When intact cells from each strain were treated with trypsin, only B. burgdorferi P66 was trypsin sensitive, indicating that K487 is surface expos...
In urban India, now a day's large number of multi story RC framed residential buildings are const... more In urban India, now a day's large number of multi story RC framed residential buildings are constructing with different ideas like soft storey or without soft story. The present paper investigates about the performance of non linear analysis of different RC framed models such as bare frame and open first storey and without soft storey. A soft story is an apartment building of three or more story located over a ground level with large openings such as parking area and soft storey is also called as open first storey. The study is carried out on G+4 storey buildings were considered as residential building which is located on zone III and the push over analysis and time history analysis are performed in the present study using sap 2000 v15.1.0 software. User defined hinge are assigned for beams and columns. Seismic performances of the frames were compared and effects of using different models were investigated. Results show a general changing pattern in performance point, capacity curve, demand spectrum curve.
We previously identified T. pallidum repeat proteins TprC/D, TprF, and TprI as candidate outer me... more We previously identified T. pallidum repeat proteins TprC/D, TprF, and TprI as candidate outer membrane proteins (OMPs) and subsequently demonstrated that TprC is not only a rare OMP but also forms trimers and has porin activity. We also reported that TprC contains N- and C-terminal domains (TprCN and TprCC) orthologous to regions in the major outer sheath protein (MOSPN and MOSPC) of T. denticola and that TprCC is solely responsible for β-barrel formation, trimerization, and porin function by the full-length protein. Herein, we show that TprI also possesses bipartite architecture, trimeric structure, and porin function and that the MOSPC-like domains of native TprC and TprI are surface-exposed in T. pallidum while their MOSPN-like domains are tethered within the periplasm. TprF, which does not contain a MOSPC-like domain, lacks amphiphilicity and porin activity, adopts an extended inflexible structure, and in T. pallidum, is tightly bound to the protoplasmic cylinder. By thermal de...
Definitive identification of Treponema pallidum (Tp) rare outer membrane proteins (OMPs) has long... more Definitive identification of Treponema pallidum (Tp) rare outer membrane proteins (OMPs) has long eluded researchers. TP0326, the sole protein in Tp with sequence homology to a Gramnegative OMP, belongs to the BamA family of proteins essential for OM biogenesis. Structural modeling predicted that five polypeptide transport-associated (POTRA) domains comprise the Nterminus of TP0326, while the C-terminus forms an 18-stranded amphipathic β-barrel. Circular dichroism, heat-modifiability by SDS-PAGE, Triton X-114 phase partitioning and liposome incorporation supported these topological predictions and confirmed that the β-barrel is responsible for the native protein's amphiphilicity. Expression analyses revealed that native TP0326 is expressed at low abundance, while a protease-surface accessibility assay confirmed surface exposure. Size-exclusion chromatography and blue native polyacrylamide gel electrophoresis revealed a modular Bam complex in Tp considerably larger than that of E. coli. Non-orthologous ancillary factors and self-association of TP0326 via its β-barrel may both contribute to the Bam complex. Tp-infected rabbits mount a vigorous antibody response to both POTRA and β-barrel portions of TP0326, whereas humans with secondary syphilis respond predominantly to POTRA. The syphilis spirochete appears to have devised a stratagem for harnessing the Bam pathway while satisfying its need to limit surface antigenicity.
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Papers by arvind anand