Structural characterization of the cell wall binding domains of Clostridium difficile toxins A and B;: Evidence that Ca2+ plays a role in toxin A cell surface association

被引:15
作者
Demarest, SJ [1 ]
Salbato, J [1 ]
Elia, M [1 ]
Zhong, JP [1 ]
Morrow, T [1 ]
Holland, T [1 ]
Kline, K [1 ]
Woodnutt, G [1 ]
Kimmel, BE [1 ]
Hansen, G [1 ]
机构
[1] Diversa Corp, Dept Prot Therapeut, San Diego, CA 92121 USA
关键词
Clostridium difficile; cell wall binding domain; protein folding; membrane binding mechanism; toxin A and B;
D O I
10.1016/j.jmb.2004.12.059
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Clostridium difficile (C. difficile) is a nosocomially acquired intestinal bacillus which can cause chronic diarrhea and life-threatening colitis. The pathogenic effects of the bacillus are mediated by the release of two toxins, A and B. The C-terminal portions of both toxins are composed of 20 and 30 residue repeats known as cell wall binding (CWB) domains. We have cloned and expressed the CWB-domains of toxins A and B and several truncated CWB-domain constructs to investigate their structure and function. The smallest CWB-domain that folded in a cooperative manner was an 11 repeat construct of toxin A. This differentiates the C-terminal domains of toxins A and B from the CWB-domain of Streptococcus pneumoniae LytA, which only requires six repeats to fold. The 11 repeat toxin A construct bound Ca2+ directly with millimolar affinity and interacted with mammalian cell surfaces in a concentration and Ca2+-dependent fashion. Millimolar Ca2+ levels also accelerated toxin mediated CHO cell killing in an in vitro cell assay. Together, the data suggest a role for extracellular Ca2+ in the sensitization of toxin A/cell-surface interactions. (C) 2005 Elsevier Ltd. All rights reserved.
引用
收藏
页码:1197 / 1206
页数:10
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