Neutralizing Antibodies Against Allosteric Proteins: Insights From a Bacterial Adhesin

Sokurenko, Evgeni V.; Tchesnokova, Veronika; Interlandi, Gianluca; Klevit, Rachel E.; Thomas, Wendy E.

doi:10.1016/j.jmb.2022.167717

Cited by 8 publications

(7 citation statements)

References 96 publications

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“…Furthermore, evidence suggests that SylH3's epitope is localized in or in proximity to subdomain 1β, not subdomains 1α or 2γ (23). With that in mind, it is possible that SylH3 has allosteric effects on RTB similar to those reported by mAbs against the bacterial mannose-specific adhesin, FimH (34).…”

Section: Introductionmentioning

confidence: 81%

An ancillary carbohydrate recognition domain on ricin toxin's B subunit is the target of receptor-blocking, neutralizing monoclonal antibodies

Mantis

Vance

Rudolph

et al. 2023

Preprint

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Monoclonal antibodies, JB4 and SylH3, neutralize ricin toxin (RT) by inhibiting the galactose-specific lectin activity of the toxin’s B subunit (RTB), which is required for cell attachment and entry. It is not immediately apparent how the antibodies accomplish this feat, considering that RTB consists of two globular domains (D1, D2) each divided into three homologous sub-domains (a, b, g) with the two functional galactosyl-specific carbohydrate recognition domains (CRDs) situated on opposite poles (sub-domains 1a and 2g). Here we report the X-ray crystal structures of JB4 and SylH3 Fab fragments bound to RTB in the context of RT. The structures revealed that neither Fab obstructed or induced detectable conformational alterations in subdomains 1a or 2g. Rather, JB4 and SylH3 Fabs recognize nearly identical epitopes within an ancillary carbohydrate recognition pocket located in subdomain 1β. Despite limited amino acid sequence similarity between SylH3 and JB4 Fabs, each paratope inserts a Phe side chain from heavy (H) chain complementarity determining region (CDR3) into the 1β CRD pocket, resulting in local aromatic stacking interactions that potentially mimic a ligand interaction. Reconciling the fact that stoichiometric amounts of SylH3 and JB4 are sufficient to disarm RTB’s lectin activity without evidence of allostery, we propose that subdomain 1β functions as a “coreceptor” required to stabilize glycan interactions principally mediated by subdomains 1a and 2g. Further investigation into subdomain 1β will yield fundamental insights into the large family of R-type lectins and open novel avenues for countermeasures aimed at preventing toxin uptake into vulnerable tissues and cells.

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Section: Introductionmentioning

confidence: 81%

An ancillary carbohydrate recognition domain on ricin toxin's B subunit is the target of receptor-blocking, neutralizing monoclonal antibodies

Mantis

Vance

Rudolph

et al. 2023

Preprint

View full text Add to dashboard Cite

show abstract

“…23 With that in mind, it is possible that SylH3 has allosteric effects on RTB similar to those reported by mAbs against the bacterial mannose-specific adhesin, FimH. 34 We therefore embarked on a study to define the molecular interactions between RT and SylH3 and JB4. As noted above, SylH3 and JB4 have been shown to neutralize RT in a wide range of cell lines and primary cells, including macrophages and lung-derived epithelial cells.…”

mentioning

confidence: 85%

“…For many toxins such as Shiga, anthrax, and Clostridium difficile B (TcdB), inhibition of attachment to host cell receptors involves antibody occupancy or occlusion of receptor binding pockets. − This is unlikely to be the case with SylH3, as RTB’s two principal carbohydrate recognition domains (CRDs) are antigenically distinct and spaced too far apart (∼50 Å) to be obstructed by a single mAb or Fab. ,, Furthermore, evidence suggests that SylH3’s epitope is localized in or in proximity to subdomain 1β, not subdomains 1α or 2γ . With that in mind, it is possible that SylH3 has allosteric effects on RTB similar to those reported by mAbs against the bacterial mannose-specific adhesin, FimH …”

mentioning

confidence: 99%

Structural Basis of Antibody-Mediated Inhibition of Ricin Toxin Attachment to Host Cells

Vance,

Rudolph,

Davis

et al. 2023

Biochemistry

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Monoclonal antibodies, JB4 and SylH3, neutralize ricin toxin (RT) by inhibiting the galactose-specific lectin activity of the B subunit of the toxin (RTB), which is required for cell attachment and entry. It is not immediately apparent how the antibodies accomplish this feat, considering that RTB consists of two globular domains (D1, D2) each divided into three homologous subdomains (α, β, γ) with the two functional galactosyl-specific carbohydrate recognition domains (CRDs) situated on opposite poles (subdomains 1α and 2γ). Here, we report the X-ray crystal structures of JB4 and SylH3 Fab fragments bound to RTB in the context of RT. The structures revealed that neither Fab obstructed nor induced detectable conformational alterations in subdomains 1α or 2γ. Rather, JB4 and SylH3 Fabs recognize nearly identical epitopes within an ancillary carbohydrate recognition pocket located in subdomain 1β. Despite limited amino acid sequence similarity between SylH3 and JB4 Fabs, each paratope inserts a Phe side chain from the heavy (H) chain complementarity determining region (CDR3) into the 1β CRD pocket, resulting in local aromatic stacking interactions that potentially mimic a ligand interaction. Reconciling the fact that stoichiometric amounts of SylH3 and JB4 are sufficient to disarm RTB’s lectin activity without evidence of allostery, we propose that subdomain 1β functions as a “coreceptor” required to stabilize glycan interactions principally mediated by subdomains 1α and 2γ. Further investigation into subdomain 1β will yield fundamental insights into the large family of R-type lectins and open novel avenues for countermeasures aimed at preventing toxin uptake into vulnerable tissues and cells.

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“…This has been especially true for Escherichia coli , one of the most common human pathogens, which causes a majority of diarrhea cases and urinary tract infections (UTIs) (including Foley catheter-associated UTIs) [ 37 , 38 , 39 ], as well as bloodstream infections [ 40 , 41 ], pneumonia in mechanically ventilated patients [ 42 ], and other extra-intestinal infections [ 43 ]. E. coli were the first bacteria for which flow-dependent shear-enhanced adhesion was demonstrated and its molecular mechanism was studied in great detail [ 7 , 29 , 44 , 45 , 46 , 47 ]. The most common adhesin of E. coli , the type 1 fimbrial adhesin FimH, has been a target for antiadhesive therapy to prevent urinary tract infections [ 7 , 47 ], and mediates shear-enhanced adhesion [ 29 , 44 , 45 , 46 ].…”

Section: Introductionmentioning

confidence: 99%

“…E. coli were the first bacteria for which flow-dependent shear-enhanced adhesion was demonstrated and its molecular mechanism was studied in great detail [ 7 , 29 , 44 , 45 , 46 , 47 ]. The most common adhesin of E. coli , the type 1 fimbrial adhesin FimH, has been a target for antiadhesive therapy to prevent urinary tract infections [ 7 , 47 ], and mediates shear-enhanced adhesion [ 29 , 44 , 45 , 46 ]. FimH binds mannosylated receptors via a catch bond mechanism, where the strength of binding to mannose is allosterically enhanced by the application of tensile force that induces separation between the adhesive and fimbria-anchoring domains of FimH E. coli [ 7 , 37 ].…”

Section: Introductionmentioning

confidence: 99%

Effect of Gravity on Bacterial Adhesion to Heterogeneous Surfaces

et al. 2023

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Bacterial adhesion is the first step in the formation of surface biofilms. The number of bacteria that bind to a surface from the solution depends on how many bacteria can reach the surface (bacterial transport) and the strength of interactions between bacterial adhesins and surface receptors (adhesivity). By using microfluidic channels and video microscopy as well as computational simulations, we investigated how the interplay between bacterial transport and adhesivity affects the number of the common human pathogen Escherichia coli that bind to heterogeneous surfaces with different receptor densities. We determined that gravitational sedimentation causes bacteria to concentrate at the lower surface over time as fluid moves over a non-adhesive region, so bacteria preferentially adhere to adhesive regions on the lower, inflow-proximal areas that are downstream of non-adhesive regions within the entered compartments. Also, initial bacterial attachment to an adhesive region of a heterogeneous lower surface may be inhibited by shear due to mass transport effects alone rather than shear forces per se, because higher shear washes out the sedimented bacteria. We also provide a conceptual framework and theory that predict the impact of sedimentation on adhesion between and within adhesive regions in flow, where bacteria would likely bind both in vitro and in vivo, and how to normalize the bacterial binding level under experimental set-ups based on the flow compartment configuration.

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Neutralizing Antibodies Against Allosteric Proteins: Insights From a Bacterial Adhesin

Cited by 8 publications

References 96 publications

An ancillary carbohydrate recognition domain on ricin toxin's B subunit is the target of receptor-blocking, neutralizing monoclonal antibodies

An ancillary carbohydrate recognition domain on ricin toxin's B subunit is the target of receptor-blocking, neutralizing monoclonal antibodies

Structural Basis of Antibody-Mediated Inhibition of Ricin Toxin Attachment to Host Cells

Effect of Gravity on Bacterial Adhesion to Heterogeneous Surfaces

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