The impact of TcdB conformational dynamics on vaccination against Clostridioides difficile

Clostridioides difficile TcdB is a multidomain ~270 kDa protein that adopts at least two distinct structural conformations influencing receptor binding, sensitivity to bile salts, and antibody reactivity. Data from the current study suggest TcdB may benefit from conformational flexibility by avoiding neutralization by vaccine induced antibodies. Experiments further demonstrate that targeting a single TcdB conformation overcomes the toxin’s ability to evade antibody neutralization. In the first series of experiments, mice were administered TcdB engineered with a 19 amino acid deletion and two site specific mutations that, in combination, brought toxicity to undetectable levels. This TcdB formulation drove robust antibody and recall responses that protected mice from weight loss but not damage to the intestines. Though all the treated mice generated similar anti-TcdB titers, only about half of the mice produced detectable neutralizing antibodies to TcdB’s open conformation. In the second part of this study, we sought to overcome this limitation to vaccine effectiveness by designing and testing an inactive form of TcdB modified to adopt only the open conformation. Treatment with this constitutively open form of TcdB resulted in an effective neutralizing antibody response in all the experimental mice. Vaccination reduced weight loss, preserved colon length and reduced selected measures of inflammatory-cell infiltration. These findings provide structure-based insights into the rational design of next generation vaccines that overcome TcdB’s conformational dynamics.