Masking autoprocessing of Clostridium difficile toxin A by the C-terminus combined repetitive oligo peptides.
Clostridium difficile toxin A and B (TcdA and TcdB) are the major virulence factors of the bacterium, both of which consist of two enzymatic domains: an effector glucosyltransferase domain (GTD) and a cysteine protease domain (CPD) responsible for autocleavage and release of GTD. Although the CPDs from both toxins share a similar structure and mechanism of hexakisphosphate (InsP6)-induced activation, TcdA is substantially less sensitive to the autocleavage as compared with TcdB. In this study, we provided evidence of inter-domain regulation of CPD activity of TcdA and its autoprocessing. The C-terminus combined repetitive oligo peptides (CROPs) of TcdA reduced the accessibility of TcdB CPD to its substrate in a chimeric toxin TxB-Ar, consequently blocking autoprocessing. Moreover, interference of antibodies with the CROPs of full-length TcdA efficiently enhanced its GTD release. In conclusion, by utilizing chimeric toxins and specific antibodies, we identified that the CROPs of TcdA plays a crucial role in controlling the InsP6-mediated activation of CPD and autocleavage of GTD. Our data provides insights on the molecular mode of action of the C. difficile toxins.
Department of Microbial Pathogenesis, University of Maryland Dental School, Baltimore, MD 21201, USA.
Yongrong
Y
Zhang
Department of Microbial Pathogenesis, University of Maryland Dental School, Baltimore, MD 21201, USA.
Therwa
T
Hamza
Department of Microbial Pathogenesis, University of Maryland Dental School, Baltimore, MD 21201, USA.
Si
S
Gao
Department of Microbial Pathogenesis, University of Maryland Dental School, Baltimore, MD 21201, USA. Electronic address: hfeng@umaryland.edu.
Hanping
H
Feng
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism / Q000633:toxicity; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism* / Q000633:toxicity; D002460:Cell Line; D002522:Chlorocebus aethiops; D016360:Clostridioides difficile / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity; D004768:Enterotoxins / Q000235:genetics / Q000378:metabolism* / Q000633:toxicity; D051379:Mice; D009842:Oligopeptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010833:Phytic Acid / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014709:Vero Cells; D014774:Virulence
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004768:Enterotoxins; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; C041348:tcdA protein, Clostridium difficile; C031815:toxB protein, Clostridium difficile; D010833:Phytic Acid
Autoprocessing; C-terminus combined repetitive oligo peptides (CROPs); Clostridium difficile; Cysteine protease domain (CPD); Toxin A