A Monoclonal Antibody against the C-Terminal Domain of Bacillus cereus Hemolysin II Inhibits HlyII Cytolytic Activity

Abstract

Bacillus cereus is the fourth most common cause of foodborne illnesses that produces a variety of pore-forming proteins as the main pathogenic factors. B. cereus hemolysin II (HlyII), belonging to pore-forming β-barrel toxins, has a C-terminal extension of 94 amino acid residues designated as HlyIICTD. An analysis of a panel of monoclonal antibodies to the recombinant HlyIICTD protein revealed the ability of the antibody HlyIIC-20 to inhibit HlyII hemolysis. A conformational epitope recognized by HlyIIC-20 was found. by the method of peptide phage display and found that it is localized in the N-terminal part of HlyIICTD. The HlyIIC-20 interacted with a monomeric form of HlyII, thus suppressing maturation of the HlyII toxin. Protection efficiencies of various B. cereus strains against HlyII were different and depended on the epitope amino acid composition, as well as, insignificantly, on downstream amino acids. Substitution of L324P and P324L in the hemolysins ATCC14579T and B771, respectively, determined the role of leucine localized to the epitope in suppressing the hemolysis by the antibody. Pre-incubation of HlyIIC-20 with HlyII prevented the death of mice up to an equimolar ratio. A strategy of detecting and neutralizing the toxic activity of HlyII could provide a tool for monitoring and reducing B. cereus pathogenicity.

A High-Homology Region in Bacterial β-Barrel Pore-Forming Toxins

“…One of them is located in the GP-5 recognize region sequence instead of Tre217 in HlyII and Ser236 in CytK2, while CytK2 and HlyIIΔCTD each have one proline residue outside this region. We have previously demonstrated the role of proline in the accessibility of mAbs amino acid sequences [10,22]. The presence of proline residues can significantly change the 3D structure of proteins and can modulate the suppression of PFTs activity.…”

“…One of the effective ways to suppress cytolytic activity is to use monoclonal antibodies (mAbs) against certain regions of the PFTs' amino acid sequences [7]. This has been demonstrated for Staphylococcus aureus α-toxin (Hla) [8], Streptococcus pneumoniae pneumolysin [9], B. cereus HlyII [10] and some others. The aim of this work was to obtain mAbs against homologous peptide of B. cereus HlyII as a tool for simultaneous identification and assessment of the level of expression of various β-PFTs.…”

Biological Efficacy of Compounds from Stingless Honey and Sting Honey against Two Pathogenic Bacteria: An In Vitro and In Silico Study

“…Hemolysin II, the target protein of the bacterium Bacillus cereus , is known to be harmful. N. Rudenko et al denoted that monoclonal antibody HlyIIC-20 has the ability to suppress HlyII hemolysis (6d5z) [ 66 , 67 ]. Therefore, this novel protein might be an antibiotic target because its pore-forming activity causes apoptosis [ 68 ].…”

Antibodies as Biosensors’ Key Components: State-of-the-Art in Russia 2020–2021

“…The mAbs described in [52] bind to two different epitopes of G protein of the respiratory syncytial virus, possess a virus-neutralizing activity, demonstrate an additive effect in interaction, detect the viral antigen in infected cell cultures, and can be used to reveal the virus in clinical material by immunofluorescence assay and ELISA. Studies using mAbs have shown the significance of the C-terminal domain of hemolysin II for the hemolytic activity of Bacillus cereus (which causes infections in persons with weakened immunity and is one of the widespread causes of intra-hospital infections) [53], as well as its ability to bind to cell membranes [54] and be oligomerized in the presence of cell membranes [55].…”