The toxic feature of two disparate pore-forming toxins, Cry delta-endotoxins from Bacillus thuringiensis (biopesticide) and CyaA-hemolysin (CyaA-Hly) from Bordetella pertussis (human pathogen causing whooping cough), is generally attributed to their capability to form oligomeric pores, causing target cell lysis. Attempts via multidisciplinary research have been made to provide more critical insights into membrane-pore formation and receptor recognition for both types of toxins. For the Cry4Ba mosquito-active toxin [1], two direct rendering techniques, single particle negative-stain EM and high-speed AFM, were employed to demonstrate a membrane-induced state of toxin monomers needed for the formation of a potential pre-pore trimer [2]. Moreover, polarity of the Cry4Ba alpha 4-alpha 5 loop residue—Asn166 was found to be important for ion permeation and pore-opening [3]. Furthermore, structural stability of two β-hairpins within the Cry4Ba receptor-binding domain was revealed to be crucial for synergistic interactions with its alternative receptor [4]. We have also disclosed functional importance of the C-terminal domain of Cry4Ba in serving as a tight-binding anchor for lipid bilayers, indicative of its potential contribution to the toxin biotoxicity [5]. Unlike the Cry4Ba toxin, CyaA-Hly requires palmitoylation at Lys983 by CyaC-acyltransferase for activating its hemolytic activity against target erythrocytes. We also revealed that the Lys983-linked palmitoyl group is not directly involved in either binding to erythrocyte membranes or toxin-induced channel conductivity, but rather required for efficient membrane inserted-pore formation [6]. We have further demonstrated that the N-terminal hydrophobic region of CyaA-Hly is also required for functional association with CyaC-acyltransferase, and hence effective palmitoylation at Lys983 [7]. We have recently provided structural insights into preferential palmitoylation of CyaA-Hly through the CyaC nucleophile-activation dyad in substrate esterolysis [8]. Interestingly, we have successfully produced CyaA-specific humanized VH/VHH nanobodies that could have potential innovative applications in developing a novel anti-pertussis agent [9], eventually being used for the benefit of mankind as a whole.

     References: [1] Kanchanawarin & Angsuthanasombat (2018) In: Computational Design of Chemicals for the Control of Mosquitoes & Their Diseases. CRC Press, pp.191-208. [2] Sriwimol et al. (2015) J. Biol. Chem. 290:20793. [3] Juntadech et al. (2014) Biochim. Biophys. ActaBiomembranes 1838:319. [4] Kaikaew et al. (2016) Biochem. Biophys. Res. Comm. 496:698. [5]. Thammasittirong et al. (2019) Toxins 11:62. [6] Meetum et al. (2017) Biochim. Biophys. ActaBiomembranes 1859:312. [7] Raksanoh et al. (2018) Biochem. Biophys. Res. Comm. 499:862. [8] Yentongchai et al. (2020) Arch. Biochem. Biophys. 694:108615. [9] Malik et al. (2016) Toxins 8:99.

1.Sriwimol, W., Aroonkesorn, A., Sakdee, S., Kanchanawarin, C., Uchihashi, T., Ando, T. & Angsuthanasombat, C. (2015) Potential pre-pore trimer formation by the Bacillus thuringiensis mosquito-specific toxin: molecular insights into a critical prerequisite of membrane-bound monomers. J. Biol. Chem. 290: 20793.

2.Malik, A., Imtong, C., Sookrung, N., Katzenmeier, G., Chaicumpa, W. & Angsuthanasombat, C. (2016) Structural characterization of humanized nanobodies with neutralizing activity against the Bordetella pertussis CyaA-hemolysin: implications for a potential epitope of toxin- protective antigen. Toxins 8: 99.

3.Meetum, K., Imtong, C., Katzenmeier, G. & Angsuthanasombat, C. (2017) Acylation of the Bordetella pertussis CyaA-hemolysin: functional implications for efficient membrane insertion and pore formation. Biochim. Biophys. Acta – Biomembrane 1859: 312.

4.Thammasittirong, A., Imtong, C., Sriwimol, W., Sakdee, S. & Angsuthanasombat, C. (2019) The C-terminal domain (DIII) of the Bacillus thuringiensis Cry4Ba toxin functions as a moiety for membrane anchoring. Toxins 11: 62.

5.Yentongchai, M., Thamwiriyasati, N., Imtong, C., Li, H.C. & Angsuthanasombat, C. (2020) Preferential modification of CyaA-hemolysin by CyaC-acyltransferase through the catalytic Ser30-His33 dyad in esterolysis of palmitoyl-donor substrate devoid of acyl carrier proteins. Arch. Biochem. Biophys. 694 (15):108615.