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Insights into the potential function and membrane organization of the TP0435 (Tp17) lipoprotein from Treponema pallidum derived from structural and biophysical analyses.

ABSTRACT: The sexually transmitted disease syphilis is caused by the bacterial spirochete Treponema pallidum. This microorganism is genetically intractable, accounting for the large number of putative and undercharacterized members of the pathogen's proteome. In an effort to ascribe a function(s) to the TP0435 (Tp17) lipoprotein, we engineered a soluble variant of the protein (rTP0435) and determined its crystal structure at a resolution of 2.42 Å. The structure is characterized by an eight-stranded ?-barrel protein with a shallow "basin" at one end of the barrel and an ?-helix stacked on the opposite end. Furthermore, there is a disulfide-linked dimer of the protein in the asymmetric unit of the crystals. Solution hydrodynamic experiments established that purified rTP0435 is monomeric, but specifically forms the disulfide-stabilized dimer observed in the crystal structure. The data herein, when considered with previous work on TP0435, imply plausible roles for the protein in either ligand binding, treponemal membrane architecture, and/or pathogenesis.

SUBMITTER: Brautigam CA

PROVIDER: S-EPMC4282407 | biostudies-literature | 2015 Jan

REPOSITORIES: biostudies-literature

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Insights into the potential function and membrane organization of the TP0435 (Tp17) lipoprotein from Treponema pallidum derived from structural and biophysical analyses.

Brautigam Chad A CA Deka Ranjit K RK Liu Wei Z WZ Norgard Michael V MV

Protein science : a publication of the Protein Society 20141025 1

The sexually transmitted disease syphilis is caused by the bacterial spirochete Treponema pallidum. This microorganism is genetically intractable, accounting for the large number of putative and undercharacterized members of the pathogen's proteome. In an effort to ascribe a function(s) to the TP0435 (Tp17) lipoprotein, we engineered a soluble variant of the protein (rTP0435) and determined its crystal structure at a resolution of 2.42 Å. The structure is characterized by an eight-stranded β-bar ...[more]

PMID: 25287511

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Project description:BACKGROUND:Syphilis continues to be a major global health threat with 11 million new infections each year, and a global burden of 36 million cases. The causative agent of syphilis, Treponema pallidum subspecies pallidum, is a highly virulent bacterium, however the molecular mechanisms underlying T. pallidum pathogenesis remain to be definitively identified. This is due to the fact that T. pallidum is currently uncultivatable, inherently fragile and thus difficult to work with, and phylogenetically distinct with no conventional virulence factor homologs found in other pathogens. In fact, approximately 30% of its predicted protein-coding genes have no known orthologs or assigned functions. Here we employed a structural bioinformatics approach using Phyre2-based tertiary structure modeling to improve our understanding of T. pallidum protein function on a proteome-wide scale. RESULTS:Phyre2-based tertiary structure modeling generated high-confidence predictions for 80% of the T. pallidum proteome (780/978 predicted proteins). Tertiary structure modeling also inferred the same function as primary structure-based annotations from genome sequencing pipelines for 525/605 proteins (87%), which represents 54% (525/978) of all T. pallidum proteins. Of the 175 T. pallidum proteins modeled with high confidence that were not assigned functions in the previously annotated published proteome, 167 (95%) were able to be assigned predicted functions. Twenty-one of the 175 hypothetical proteins modeled with high confidence were also predicted to exhibit significant structural similarity with proteins experimentally confirmed to be required for virulence in other pathogens. CONCLUSIONS:Phyre2-based structural modeling is a powerful bioinformatics tool that has provided insight into the potential structure and function of the majority of T. pallidum proteins and helped validate the primary structure-based annotation of more than 50% of all T. pallidum proteins with high confidence. This work represents the first T. pallidum proteome-wide structural modeling study and is one of few studies to apply this approach for the functional annotation of a whole proteome.

Project description:In recent years, considerable progress has been made in topologically and functionally characterizing integral outer membrane proteins (OMPs) of Treponema pallidum subspecies pallidum, the syphilis spirochete, and identifying its surface-exposed ?-barrel domains. Extracellular loops in OMPs of Gram-negative bacteria are known to be highly variable. We examined the sequence diversity of ?-barrel-encoding regions of tprC, tprD, and bamA 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 T. pallidum 0548 (tp0548) genotyping and tp0558 sequences to assign strains to the Nichols or SS14 clades. We found that (i) ?-barrels in clinical strains could be grouped according to allelic variants in T. pallidum subsp. pallidum reference genomes; (ii) for all three OMP loci, clinical strains within the Nichols or SS14 clades often harbored ?-barrel variants that differed from the Nichols and SS14 reference strains; and (iii) OMP variable regions often reside in predicted extracellular loops containing B-cell epitopes. On the basis of structural models, nonconservative amino acid substitutions in predicted transmembrane ?-strands of T. pallidum repeat C (TprC) and TprD2 could give rise to functional differences in their porin channels. OMP profiles of some clinical strains were mosaics of different reference strains and did not correlate with results from enhanced molecular typing. Our observations suggest that human host selection pressures drive T. pallidum subsp. pallidum OMP diversity and that genetic exchange contributes to the evolutionary biology of T. pallidum subsp. pallidum They also set the stage for topology-based analysis of antibody responses to OMPs and help frame strategies for syphilis vaccine development.IMPORTANCE Despite recent progress characterizing outer membrane proteins (OMPs) of Treponema pallidum, little is known about how their surface-exposed, ?-barrel-forming domains vary among strains circulating within high-risk populations. In this study, sequences for the ?-barrel-encoding regions of three OMP loci, tprC, tprD, and bamA, in T. pallidum subsp. pallidum isolates from a large number of patient specimens from geographically disparate sites were examined. Structural models predict that sequence variation within ?-barrel domains occurs predominantly within predicted extracellular loops. Amino acid substitutions in predicted transmembrane strands that could potentially affect porin channel function were also noted. Our findings suggest that selection pressures exerted within human populations drive T. pallidum subsp. pallidum OMP diversity and that recombination at OMP loci contributes to the evolutionary biology of syphilis spirochetes. These results also set the stage for topology-based analysis of antibody responses that promote clearance of T. pallidum subsp. pallidum and frame strategies for vaccine development based upon conserved OMP extracellular loops.

Dataset Information

Insights into the potential function and membrane organization of the TP0435 (Tp17) lipoprotein from Treponema pallidum derived from structural and biophysical analyses.

Publications

Insights into the potential function and membrane organization of the TP0435 (Tp17) lipoprotein from Treponema pallidum derived from structural and biophysical analyses.

Similar Datasets

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