Abstract
Several pathogenic bacteria are able to induce the attaching and effacing (A/E) lesion. The A/E lesion is caused by effector proteins, such as Escherichia coli-secreted protein B (EspB), responsible together with Escherichia coli-secreted protein D for forming a pore structure on the host cell, which allows the translocation of effector proteins. Different variants of this protein can be found in E. coli strains, and during natural infection or when this protein is injected, this leads to variant-specific production of antibodies, which may not be able to recognize other variants of this bacterial protein. Herein, we describe the production of a hybrid recombinant EspB toxin that comprises all known variants of this protein. This recombinant protein could be useful as an antigen for the production of antibodies with broad-range detection of EspB-bearing bacteria, or as an antigen that could be used in vaccine formulation to generate antibodies against different EspB variants, thereby increasing immunization potential. In addition, the recombinant protein allowed us to analyze its secondary structure, to propose the immunogenic regions of EspB variants, and also to characterize anti-EspB antibodies. Our results suggest that this hybrid protein or a protein composed of the conserved immunogenic regions could be used for a variety of clinical applications.
Highlights
Gram-negative pathogenic bacteria, such as enteropathogenic Escherichia coli (EPEC), enterohemorrhagic Escherichia coli (EHEC), and Citrobacter rodentium are able to induce attaching and effacing (A/E) lesion [1,2,3]
The hybrid recombinant EspB (rEspB) protein was obtained from the E. coli BL21 transformed with a plasmid harboring the hybrid espB gene
Escherichia coli-secreted protein B protein is translocated into the host cell through a T3SS and together with Escherichia coli-secreted protein D (EspD) is responsible for assembling a multimeric pore in the eukaryotic membrane, contributing to the hallmarks of the A/E lesion
Summary
Gram-negative pathogenic bacteria, such as enteropathogenic Escherichia coli (EPEC), enterohemorrhagic Escherichia coli (EHEC), and Citrobacter rodentium are able to induce attaching and effacing (A/E) lesion [1,2,3]. The Esp responsible for the syringe-like structure of T3SS is secreted protein A (EspA), which is the needle-shaped protein of approximately 25 kDa, while secreted proteins B [Escherichia coli-secreted protein B (EspB)] and D [Escherichia coli-secreted protein D (EspD)] are responsible for the pore structure assembled in the eukaryotic membrane [8]. Eliciting antibodies against bacterial colonization factors have been proposed as a vaccination strategy to prevent pathogenic E. coli infection [18]. Antibodies against the T3SS proteins, such as EspA, EspB, and EspD, have been detected in the serum from patients with diarrheagenic E. coli infections, demonstrating their immunogenic potential [19,20,21,22]. We synthetically constructed a hybrid recombinant EspB (rEspB), representative of all known variants to date, and characterized its secondary structure, which allowed us to propose an immunogenic domain
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