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Structural and functional analysis of the CspB protease required for Clostridium spore germination
- Source :
- PLoS Pathogens, Vol 9, Iss 2, p e1003165 (2013), PLoS Pathogens
- Publication Year :
- 2013
- Publisher :
- Public Library of Science (PLoS), 2013.
-
Abstract
- Spores are the major transmissive form of the nosocomial pathogen Clostridium difficile, a leading cause of healthcare-associated diarrhea worldwide. Successful transmission of C. difficile requires that its hardy, resistant spores germinate into vegetative cells in the gastrointestinal tract. A critical step during this process is the degradation of the spore cortex, a thick layer of peptidoglycan surrounding the spore core. In Clostridium sp., cortex degradation depends on the proteolytic activation of the cortex hydrolase, SleC. Previous studies have implicated Csps as being necessary for SleC cleavage during germination; however, their mechanism of action has remained poorly characterized. In this study, we demonstrate that CspB is a subtilisin-like serine protease whose activity is essential for efficient SleC cleavage and C. difficile spore germination. By solving the first crystal structure of a Csp family member, CspB, to 1.6 Å, we identify key structural domains within CspB. In contrast with all previously solved structures of prokaryotic subtilases, the CspB prodomain remains tightly bound to the wildtype subtilase domain and sterically occludes a catalytically competent active site. The structure, combined with biochemical and genetic analyses, reveals that Csp proteases contain a unique jellyroll domain insertion critical for stabilizing the protease in vitro and in C. difficile. Collectively, our study provides the first molecular insight into CspB activity and function. These studies may inform the development of inhibitors that can prevent clostridial spore germination and thus disease transmission.<br />Author Summary Clostridium difficile is the leading cause of health-care associated diarrhea worldwide. C. difficile infections begin when its spores transform into vegetative cells during a process called germination. In Clostridium sp., germination requires that the spore cortex, a thick, protective layer, be removed by the cortex hydrolase SleC. While previous studies have shown that SleC activity depends on a subtilisin-like protease, CspB, the mechanisms regulating CspB function have not been characterized. In this study, we solved the first crystal structure of the Csp family of proteases and identified its key functional regions. We determined that CspB carries a unique jellyroll domain required for stabilizing the protein both in vitro and in C. difficile and a prodomain required for proper folding of the protease. Unlike all other prokaryotic subtilisin-like proteases, the prodomain remains bound to CspB and inhibits its protease activity until the germination signal is sensed. Our study provides new insight into how germination is regulated in C. difficile and may inform the development of inhibitors that can prevent germination and thus C. difficile transmission.
- Subjects :
- Models, Molecular
Proteases
Clostridium perfringens
QH301-705.5
medicine.medical_treatment
Immunology
Peptidoglycan
Biochemistry
Microbiology
Subtilase
03 medical and health sciences
chemistry.chemical_compound
Clostridium
Bacterial Proteins
Catalytic Domain
Virology
Genetics
medicine
Spore germination
Humans
Biology (General)
Biology
Molecular Biology
030304 developmental biology
Spores, Bacterial
Serine protease
0303 health sciences
Protease
biology
Clostridioides difficile
030306 microbiology
fungi
Subtilisin
RC581-607
biology.organism_classification
Protein Structure, Tertiary
chemistry
Clostridium Infections
biology.protein
Parasitology
Serine Proteases
Immunologic diseases. Allergy
Research Article
Subjects
Details
- Language :
- English
- ISSN :
- 15537374 and 15537366
- Volume :
- 9
- Issue :
- 2
- Database :
- OpenAIRE
- Journal :
- PLoS Pathogens
- Accession number :
- edsair.doi.dedup.....0d5e726dbfbb3b0afa9d72466973c7e4