*1.C.73 The Pseudomonas* Exotoxin A (P-ExoA) Family** Exposure to low endosomal pH during internalization of Pseudomonas exotoxin A (PE) triggers membrane insertion of its translocation domain. This process is a prerequisite for PE translocation to the cytosol where it inactivates protein synthesis. Although hydrophobic helices enable membrane insertion of related bacterial toxins such as diphtheria toxin, the PE translocation domain is devoid of hydrophobic stretches. The structural features triggering acid-induced membrane insertion of PE have recently been elucidated (Méré et al., 2005). At neutral pH, a Trp is buried in a hydrophobic pocket, closed by the smallest α-helix of the translocation domain. Upon acidification, protonation of the Asp that is the N-cap residue of the helix leads to its destabilization, enabling Trp side chain insertion into the endosome membrane (Méré et al., 2005). This tryptophan-based membrane insertion system is similar to the membrane-anchoring mechanism of human annexin-V. P-ExoA is 613 aas long and consists of 3 structural/functional domains. Domain I binds to the α₂-macroglobulin/low density lipoprotein receptor-related protein, enabling internalization via receptor-mediated endocytosis. Domain II then mediates translocation into the cytosol of the entire toxin or of a carboxyl-terminal fragment generated by furin proteolysis and encompassing domain III and most of domain II. Finally, domain III catalyzes the ADP ribosylation of elongation factor 2, thereby inhibiting protein synthesis and killing the cell. P-ExoA has only one homologue in the current (7/05) NCBI database, a hypothetical exotoxin A from Vibrio cholerae. It is 666 aas long and exhibits 33% identity with P-ExoA throughout almost all of its length. A C-terminal domain (residues 436-502) shows significant sequence similarity (27% identity) with a region (42-167) of diphtheria toxin from C. diphtheriae (TC #1.C.7.1.1) as well as a region (116-239) with 22% identity with a region (314-420) of a putative exported protein from Yersinia species such as Y. pestis (806 aas; AAS62938).
References:
Basso, P., M. Ragno, S. Elsen, E. Reboud, G. Golovkine, S. Bouillot, P. Huber, S. Lory, E. Faudry, and I. Attrée. (2017). Pseudomonas aeruginosa Pore-Forming Exolysin and Type IV Pili Cooperate To Induce Host Cell Lysis. MBio 8:.
Huber, P. (2022). ExlA: A New Contributor to Virulence. Front Cell Infect Microbiol 12: 929150.
Lugo MR. and Merrill AR. (2015). A comparative structure-function analysis of active-site inhibitors of Vibrio cholerae cholix toxin. J Mol Recognit. 28(9):539-52.
Méré, J., J. Morlon-Guyot, A. Bonhoure, L. Chiche, and B. Beaumelle. (2005). Acid-triggered membrane insertion of Pseudomonas exotoxin A involves an original mechanism based on pH-regulated tryptophan exposure. J. Biol. Chem. 280: 21194-21201.
Rasper, D.M. and A.R. Merrill. (1994). Evidence for the modulation of Pseudomonas aeruginosa exotoxin A-induced pore formation by membrane surface charge density. Biochemistry 33: 12981-12989.
Reboud, E., S. Bouillot, S. Patot, B. Béganton, I. Attrée, and P. Huber. (2017). Pseudomonas aeruginosa ExlA and Serratia marcescens ShlA trigger cadherin cleavage by promoting calcium influx and ADAM10 activation. PLoS Pathog 13: e1006579. [Epub: Ahead of Print]
Zalman, L.S. and B.J. Wisnieski. (1985). Characterization of the insertion of Pseudomonas exotoxin A into membranes. Infect. Immun. 50: 630-635.
Examples:
TC#	Name	Organismal Type	Example
1.C.73.1.1	Pore-forming exotoxin A (chain A; ExlA) (Rasper and Merrill 1994; Méré et al., 2005). Pore-formation has been demonstrated (Zalman and Wisnieski 1985). Secretion depends on ExlB, a Two Partner Secretion (TPS; TC# 1.B.20) system, as well as type IV pili. The protein has three domains: an N-terminal hemolyin domain, a central RGD motif domain, and a C-terminal domain required for cell lysis. Pore-formation precedes lysis (Basso et al. 2017). ExlA triggers cadherin cleavage by promoting calcium influx which activates ADAM10 for proteolysis (Reboud et al. 2017). ExlA possesses pore-forming activity and is cytolytic for most human cell types. It belongs to a class of poorly characterized bacterial toxins, sharing a similar protein domain organization and a common secretion pathway (Huber 2022).	Proteobacteria	*Exotoxin A (ExlA) of Pseudomonas aeruginosa* (P11439)**

1.C.73.1.2	The cholix toxin. The NAD-dependent ADP-ribosyltransferase (ADPRT) catalyzes transfer of the ADP-ribosyl moiety of oxidized NAD onto eukaryotic elongation factor 2 (eEF-2), thus arresting protein synthesis. It may use the eukaryotic pro-low-density lipoprotein receptor-related protein 1 (LRP1) to enter mouse cells, Cholix toxin shares structural and functional properties with Pseudomonas aeruginosa exotoxin A and Corynebacterium diphtheriae diphtheria toxin (Lugo and Merrill 2015).	Proteobacteria	Cholix toxin of Vibrio cholera

1.C.73.1.3	Exotoxin A of 241 aas	Proteobacteria	Exotoxin A of Cystobacter fuscus

Examples:
TC#	Name	Organismal Type	Example
1.C.73.2.1	Uncharacterzed protein of 679 aas	Proteobacteria	UP of Yersinia frederiksenii

1.C.73.2.2	Exotoxin of 806 aas	Proteobacteria	Exotoxin of Yersinia similis

Examples:
TC#	Name	Organismal Type	Example
1.C.73.3.1	Uncharacterized protein of 698 aas	Actinobacteria	UP of Mycobacterium gastri

1.C.73.3.2	Uncharacterized toxin of 736 aas.		Toxin of Chloracidobacterium thermophilum

1.C.73.3.3	Putative toxin of 937 aa	Toxin of Blastopirellula marina	Toxin of Blastopirellula marina

1.C.73.3.4	Putative toxin of 679 aas		*Toxin of Gloeocapsa* sp. PCC 7428**