Dataset Information

Structure-Immunogenicity Relationship of ?- and ?-Tetrasaccharide Glycoforms from Bacillus anthracis Exosporium and Fragments Thereof.

ABSTRACT: The tetrasaccharide (2-O-methyl-4-(3-hydroxy-3-methylbutamido)-4,6-dideoxy-?-d-glucopyranosyl-(1?3)-?-l-rhamnopyranosyl-(1?3)-?-l-rhamnopyranosyl-(1?2)-l-rhamnopyranose) from the major exosporium protein (BclA) of Bacillus anthracis has been proposed as a target for development of diagnostics and immune therapy or prophylaxis. While the immunodominant character of the anthrose residue has been previously elucidated, the role of the stereochemical configuration of the downstream rhamnose is unknown. Because the linkage of this residue to the GlcNAc bridging the glycan and the protein is lost during isolation of the tetrasaccharide, its ?- and ?-glycoforms have been synthesized. Herein, we prepared neoglycoconjugates from a series of fragments of the tetrasaccharide, including the complete ?- and ?-tetrasaccharide glycoforms, a 2-demethoxylated version of the ?-tetrasaccharide, and the ?- and ?-trirhamnosides and CRM197. By immunization of mice, we showed that the anti ?- and ?-tetrasaccharide serum equally recognized both glycoforms. In contrast the sera produced following immunization with the ?- and ?-trirhamnoside fragments exhibited higher recognition for their own antigens than for their anomeric counterparts. The anti ?- and ?-tetrasaccharide sera recognized Sterne spores in a comparable fashion. ?BclA spores not expressing the major exosporium protein were also recognized by the same sera, while mutants that produced the carbohydrate antigen with deletion of either rhamnose or anthrose were not. The tetrasaccharide could, therefore, be expressed in proteins other than BlcA. This work proves that ?- and ?-tetrasaccharide are equally potent immunogens.

SUBMITTER: De Ricco R

PROVIDER: S-EPMC6222408 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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Structure-Immunogenicity Relationship of α- and β-Tetrasaccharide Glycoforms from Bacillus anthracis Exosporium and Fragments Thereof.

De Ricco Riccardo R Ventura Christy L CL Carboni Filippo F Saksena Rina R Kováč Pavol P Adamo Roberto R

Molecules (Basel, Switzerland) 20180820 8

The tetrasaccharide (2-O-methyl-4-(3-hydroxy-3-methylbutamido)-4,6-dideoxy-α-d-glucopyranosyl-(1→3)-α-l-rhamnopyranosyl-(1→3)-α-l-rhamnopyranosyl-(1→2)-l-rhamnopyranose) from the major exosporium protein (BclA) of Bacillus anthracis has been proposed as a target for development of diagnostics and immune therapy or prophylaxis. While the immunodominant character of the anthrose residue has been previously elucidated, the role of the stereochemical configuration of the downstream rha ...[more]

PMID: 30127242

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Project description:In preliminary experiments, the absence of anthrose in our genetic Sterne knockout was found to decrease pagA expression in BHI broth containing glucose (Fig 5A and B). This contrasted with the large spike in pagA expression when the Sterne antC mutant is grown in protein rich HIB media which does not contain sugars. When exogenous anthrose was added during growth in BHI broth, expression from PpagA increased in both WT (purple line in Fig 5A) and antC Sterne (purple line in Fig 5B). Many bacteria respond to levels of metabolic intermediates and modulate gene expression accordingly. We hypothesized intracellular or extracellular anthrose levels could impact the vegetative cells through modification of gene expression during vegetative growth. Wildtype B. anthracis Sterne was grown in the presence and absence of exogenously added pure anthrose to avoid any possibility of confounding data in a deletion mutant. Bacteria were harvested at two timepoints post-addition of anthrose, 30 min and 2 h, and compared to diluent only treated cultures (Fig 5C). After 30 min treatment, there were 6 genes significantly upregulated more than two-fold while 17 genes were downregulated (Fig 5D). These genes were mainly involved in generating metabolic intermediates such as pyruvate and trehalose. The most upregulated gene was a putative membrane protein (log2FC=10.98) followed by a gene encoding a GerPF homologue at a log2FC of 10.98. Mutation of GerPF family proteins have been linked to a super-dormant spore phenotype[10]. The gene experiencing the highest downregulation was a putative lipoprotein (log2FC = -20.02). No genes located on pXO1 were detected at 30 min. A subset of differentially transcribed genes from the 30 min timepoint are listed in Fig 5F. After 2 h of anthrose treatment, 52 genes experienced significant up regulation, 18 of which were hypothetical proteins (Fig 5E). Forty-five genes experienced significant down regulation across replicates, 13 of which were hypothetical proteins.

Project description:Anthrax, caused by the bacterium Bacillus anthracis, is a disease of historical and current importance that is found throughout the world. The basis of its historical transmission is anecdotal and its true global population structure has remained largely cryptic. Seven diverse B. anthracis strains were whole-genome sequenced to identify rare single nucleotide polymorphisms (SNPs), followed by phylogenetic reconstruction of these characters onto an evolutionary model. This analysis identified SNPs that define the major clonal lineages within the species. These SNPs, in concert with 15 variable number tandem repeat (VNTR) markers, were used to subtype a collection of 1,033 B. anthracis isolates from 42 countries to create an extensive genotype data set. These analyses subdivided the isolates into three previously recognized major lineages (A, B, and C), with further subdivision into 12 clonal sub-lineages or sub-groups and, finally, 221 unique MLVA15 genotypes. This rare genomic variation was used to document the evolutionary progression of B. anthracis and to establish global patterns of diversity. Isolates in the A lineage are widely dispersed globally, whereas the B and C lineages occur on more restricted spatial scales. Molecular clock models based upon genome-wide synonymous substitutions indicate there was a massive radiation of the A lineage that occurred in the mid-Holocene (3,064-6,127 ybp). On more recent temporal scales, the global population structure of B. anthracis reflects colonial-era importation of specific genotypes from the Old World into the New World, as well as the repeated industrial importation of diverse genotypes into developed countries via spore-contaminated animal products. These findings indicate humans have played an important role in the evolution of anthrax by increasing the proliferation and dispersal of this now global disease. Finally, the value of global genotypic analysis for investigating bioterrorist-mediated outbreaks of anthrax is demonstrated.

Dataset Information

Structure-Immunogenicity Relationship of ?- and ?-Tetrasaccharide Glycoforms from Bacillus anthracis Exosporium and Fragments Thereof.

Publications

Structure-Immunogenicity Relationship of α- and β-Tetrasaccharide Glycoforms from <i>Bacillus anthracis</i> Exosporium and Fragments Thereof.

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