Publication: Bordetella pertussis CyaA-RTX subdomain requires calcium ions for structural stability against proteolytic degradation
Issued Date
2011-02-01
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ISSN
10465928
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2-s2.0-78649322168
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Mahidol University
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SCOPUS
Bibliographic Citation
Protein Expression and Purification. Vol.75, No.2 (2011), 127-132
Suggested Citation
Pichaya Pojanapotha, Niramon Thamwiriyasati, Busaba Powthongchin, Gerd Katzenmeier, Chanan Angsuthanasombat Bordetella pertussis CyaA-RTX subdomain requires calcium ions for structural stability against proteolytic degradation. Protein Expression and Purification. Vol.75, No.2 (2011), 127-132. doi:10.1016/j.pep.2010.07.012 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/11593
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Title
Bordetella pertussis CyaA-RTX subdomain requires calcium ions for structural stability against proteolytic degradation
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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. © 2010 Elsevier Inc. All rights reserved.