Structural Modeling of the Treponema pallidum Outer Membrane Protein Repertoire: a Road Map for Deconvolution of Syphilis Pathogenesis and Development of a Syphilis Vaccine

Abstract

Treponema pallidum, an obligate human pathogen, has an outer membrane (OM) whose physical properties, ultrastructure and composition differ markedly from those of phylogenetically distant Gram-negatives. We developed structural models for the outer membrane protein (OMP) repertoire of T. pallidum Nichols using solved Gram-negative structures, computational tools, and small angle X-ray scattering (SAXS) of selected recombinant periplasmic domains. The T. pallidum ‘OMPeome’ harbors two ‘stand-alone’ proteins (BamA and LptD) involved in OM biogenesis and four paralogous families involved in influx/efflux of small molecules: 8-stranded β-barrels, long-chain fatty acid transporters (FadLs), OM factors (OMFs) for efflux pumps, and T. pallidum repeat proteins (Tprs). BamA (TP0326), the central component of a BAM/TAM hybrid, possesses a highly flexible POTRA1-5 predicted to interact with TamB (TP0325). TP0515, an LptD ortholog, contains a novel, unstructured C-terminal domain that models inside the β-barrel. T. pallidum has four 8-stranded β-barrels, each containing positively charged extracellular loops that could contribute to pathogenesis. Three of five FadL-like orthologs have a novel α-helical, presumptively periplasmic C-terminal extension. SAXS and structural modeling further supported the bipartite membrane topology and tri-domain architecture of full-length members of the Tpr family. T. pallidum’s two efflux pumps presumably extrude noxious small molecules via four co-expressed OMFs with variably charged tunnels. For BamA, LptD and OMFs, we modeled the molecular machines that deliver their substrates into the OM or external milieu. The spirochete’s extended families of OM transporters collectively confer a broad capacity for nutrient uptake. The models also furnish a structural roadmap for vaccine development. Importance The unusual outer membrane (OM) of T. pallidum, the syphilis spirochete, is the ultrastructural basis for its well-recognized capacity for invasiveness, immune evasion, and persistence. In recent years, we have made considerable progress identifying T. pallidum’s repertoire of OMPs. Herein, we developed three-dimensional (3D) models for the T. pallidum Nichols OMPeome using structural modeling, bioinformatics, and solution scattering. The OM contains three families of OMP transporters, an OMP family involved in extrusion of noxious molecules, and two ‘stand alones’ involved in OM biogenesis. This work represents a major advance towards elucidating host-pathogen interactions during syphilis, understanding how T. pallidum, an extreme auxotroph, obtains a wide array of biomolecules from its obligate human host, and developing a vaccine with global efficacy.

Treponema pallidum genome sequencing from six continents reveals variability in vaccine candidate genes and dominance of Nichols clade strains in Madagascar

“…In contrast to TP0326, the structure and function of which has been studied extensively, less is known about TP0548, a predicted homolog of the E. coli fatty acid transporter FadL. We predict the structure to be a 14-stranded β-barrel, with periplasmic C-terminal α-helices, consistent with previous studies [48]. Prediction of linear B cell epitopes (BCEs) using BepiPred 2.0 [63] Both TP0966 and TP0967 are predicted to be orthologs of the E. coli efflux pump TolC [48], and are predicted to have a tri-partite structure, with each monomer contributing four βstrands to β-barrel that spans the outer membrane with BCEs predicted within the ECLs [48].…”

“…pallidum subclades are in proteins putatively located in the outer membrane, or known to react with serum from syphilis patients (Fig 4) [45,46]. These data, along with recent structural modeling of T. pallidum outer membrane proteins showing that putative B cell epitopes are primarily found on the protein surface predicted to face the host [48], strongly suggest that immune pressure is the most important driver of mutation in T. pallidum subsp. pallidum.…”

“…gonorrhoeae BamA (TP0326) and E . coli FadL (TP0548) and TolC (TP0966, TP0967) [ 48 ] and reviewed in [ 49 ]. TP0136 is a lipoprotein that appears to be localized to the outer membrane, where it functions as a fibronectin- and laminin-binding adhesin [ 50 – 52 ].…”

“…Of the six proteins that undergo mutation along four or more of the 14 branches in the phylogeny (Fig 4D and S3 Data-Heatmap), five (TP0136, TP0326, TP0548, TP0966, and TP0967) are known to be antigenic. TP0326, TP0548, TP0966, and TP0967 are likely outer membrane proteins based on their homology to N. gonorrhoeae BamA (TP0326) and E. coli FadL (TP0548) and TolC (TP0966, TP0967) [48] and reviewed in [49]. TP0136 is a lipoprotein that appears to be localized to the outer membrane, where it functions as a fibronectin-and laminin-binding adhesin [50][51][52].…”

A 2021 Update on Syphilis: Taking Stock from Pathogenesis to Vaccines

“…TP is characterized by a rare outer membrane and a slow replication rate (about 33 h) with a dynamic regulation of TP genes, which is important for its successful colonization, dissemination, and invasion in hosts [12,13]. TP repeat genes (Tpr) encode proteins that mediate attachment to host tissue and function as porins.…”