Structure-Function Analysis of Inositol Hexakisphosphate-induced Autoprocessing in Clostridium difficile Toxin A

被引:89
作者
Pruitt, Rory N. [1 ]
Chagot, Benjamin [2 ]
Cover, Michael [1 ]
Chazin, Walter J. [2 ]
Spiller, Ben [1 ,3 ]
Lacy, D. Borden [1 ,2 ]
机构
[1] Vanderbilt Univ, Med Ctr, Dept Microbiol & Immunol, Nashville, TN 37232 USA
[2] Vanderbilt Univ, Med Ctr, Dept Biochem, Nashville, TN 37232 USA
[3] Vanderbilt Univ, Med Ctr, Dept Pharmacol, Nashville, TN 37232 USA
基金
美国能源部; 美国国家卫生研究院;
关键词
CYSTEINE PROTEASE DOMAIN; RTX TOXIN; CLEAVAGE; CELLS; GLUCOSYLATION; SITE;
D O I
10.1074/jbc.M109.018929
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
The action of Clostridium difficile toxins A and B depends on inactivation of host small G-proteins by glucosylation. Cellular inositol hexakisphosphate (InsP6) induces an autocatalytic cleavage of the toxins, releasing an N-terminal glucosyltransferase domain into the host cell cytosol. We have defined the cysteine protease domain (CPD) responsible for autoprocessing within toxin A (TcdA) and report the 1.6 angstrom x-ray crystal structure of the domain bound to InsP6. InsP6 is bound in a highly basic pocket that is separated from an unusual active site by a beta-flap structure. Functional studies confirm an intramolecular mechanism of cleavage and highlight specific residues required for InsP6-induced TcdA processing. Analysis of the structural and functional data in the context of sequences from similar and diverse origins highlights a C-terminal extension and a pi-cation interaction within the beta-flap that appear to be unique among the large clostridial cytotoxins.
引用
收藏
页码:21934 / 21940
页数:7
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