title: Disulphide bond restrains the C-terminal region of thermostable direct hemolysin during folding to promote oligomerization.
abstract: Pore-forming toxins (PFTs) are typically produced as water-soluble monomers, which upon interacting with target cells assemble into transmembrane oligomeric pores. Vibrio parahaemolyticus thermostable direct hemolysin (TDH) is an atypical PFT that exists as a tetramer in solution, prior to membrane binding. The TDH structure highlights a core β-sandwich domain similar to those found in the eukaryotic actinoporin family of PFTs. However, the TDH structure harbors an extended C-terminal region (CTR) that is not documented in the actinoporins. This CTR remains tethered to the β-sandwich domain through an intra-molecular disulphide bond. Part of the CTR is positioned at the inter-protomer interface in the TDH tetramer. Here we show that the truncation, as well as mutation, of the CTR compromise tetrameric assembly, and the membrane-damaging activity of TDH. Our study also reveals that intra-protomer disulphide bond formation during the folding/assembly process of TDH restrains the CTR to mediate its participation in the formation of inter-protomer contact, thus facilitating TDH oligomerization. However, once tetramerization is achieved, disruption of the disulphide bond does not affect oligomeric assembly. Our study provides critical insights regarding the regulation of the oligomerization mechanism of TDH, which has not been previously documented in the PFT family.
authors: Centre for Protein Science, Design and Engineering, Department of Biological Sciences, Indian Institute of Science Education and Research Mohali, Sector 81, SAS Nagar, Manauli, Mohali, Punjab 140306, India.
authors: Nidhi
authors: N
authors: Kundu
authors: Centre for Protein Science, Design and Engineering, Department of Biological Sciences, Indian Institute of Science Education and Research Mohali, Sector 81, SAS Nagar, Manauli, Mohali, Punjab 140306, India.
authors: Swapnil
authors: S
authors: Tichkule
authors: Centre for Protein Science, Design and Engineering, Department of Biological Sciences, Indian Institute of Science Education and Research Mohali, Sector 81, SAS Nagar, Manauli, Mohali, Punjab 140306, India.
authors: Shashi Bhushan
authors: SB
authors: Pandit
authors: Centre for Protein Science, Design and Engineering, Department of Biological Sciences, Indian Institute of Science Education and Research Mohali, Sector 81, SAS Nagar, Manauli, Mohali, Punjab 140306, India.
authors: Kausik
authors: K
authors: Chattopadhyay
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology; D001427:Bacterial Toxins / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D001665:Binding Sites; D003001:Cloning, Molecular; D004220:Disulfides / Q000737:chemistry*; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006460:Hemolysin Proteins / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D056004:Molecular Dynamics Simulation; D009154:Mutation; D011485:Protein Binding; D000072756:Protein Conformation, alpha-Helical; D000072757:Protein Conformation, beta-Strand; D017510:Protein Folding; D054730:Protein Interaction Domains and Motifs; D055503:Protein Multimerization; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014736:Vibrio parahaemolyticus / Q000737:chemistry* / Q000472:pathogenicity
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004220:Disulfides; D006460:Hemolysin Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; C068489:thermostable direct hemolysin