Bordetella pertussis CyaA-RTX subdomain requires calcium ions for structural stability against proteolytic degradation |
| |
| Authors: | Pichaya Pojanapotha Niramon Thamwiriyasati Busaba Powthongchin Gerd Katzenmeier Chanan Angsuthanasombat |
| |
| Affiliation: | 1. Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, CB2 1GA Cambridge, United Kingdom;2. Life Sciences Group, Institut Laue-Langevin, 71 Avenue des Martyrs, CS 20156, Grenoble, Cedex 9, France;3. Faculty of Natural Sciences, Keele University, ST5 5BG Staffordshire, United Kingdom;4. Department of Genetics, University of Cambridge, Downing Street, CB2 3EH Cambridge, United Kingdom;5. Mass Spectrometry Laboratory, Department of Biophysics, Institute of Biochemistry and Biophysics, Polish Academy of Sciences, 5A Pawinskiego Street, 02-106 Warsaw, Poland;6. European Molecular Biology Laboratory, Grenoble Outstation, 71 Avenue des Martyrs, 38042 Grenoble, France;7. Unit for Virus Host–Cell Interactions, University Grenoble Alpes–European Molecular Biology Laboratory–CNRS, 71 Avenue des Martyrs, 38042 Grenoble, France |
| |
| Abstract: | Previously, the 126-kDa Bordetella pertussis CyaA pore-forming (CyaA-PF) domain expressed in Escherichia coli was shown to retain its hemolytic activity. Here, a 100-kDa RTX (Repeat-in-ToXin) subcloned fragment (CyaA-RTX) containing a number of putative calcium-binding repeats was further investigated. The recombinant CyaA-RTX protein, although expressed as a soluble form in a protease-deficient E. coli strain BL21(DE3)pLysS, was found to be highly sensitive to proteolytic degradation. Interestingly, the addition of calcium ions in a millimolar range into the CyaA-RTX preparation significantly prevented the degradation. Moreover, levels of proteolytic degradation were dependent on calcium concentrations, implying an important role for calcium-binding sites in the RTX subdomain for structural stability. Homology-based modeling of the repetitive blocks in the CyaA-RTX subdomain supports that this calcium-bound protein folds into a parallel β-roll structure with calcium ions acting as a structural stabilizing bridge. |
| |
| Keywords: | |
| 本文献已被 ScienceDirect 等数据库收录！ |
|
