Localization of the Clostridium difficile cysteine protease Cwp84 and insights into its maturation process. 2011

Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
EA 4043, Université Paris-Sud 11, Faculté de Pharmacie, Département de Microbiologie, 5 Rue Jean Baptiste Clément, 92296 Châtenay-Malabry Cedex, France.

Clostridium difficile is a nosocomial pathogen involved in antibiotic-associated diarrhea. C. difficile expresses a cysteine protease, Cwp84, which has been shown to degrade some proteins of the extracellular matrix and play a role in the maturation of the precursor of the S-layer proteins. We sought to analyze the localization and the maturation process of this protease. Two identifiable forms of the protease were found to be associated in the bacteria: a form of ∼80 kDa and a cleaved one of 47 kDa, identified as the mature protease. They were found mainly in the bacterial cell surface fractions and weakly in the extracellular fraction. The 80-kDa protein was noncovalently associated with the S-layer proteins, while the 47-kDa form was found to be tightly associated with the underlying cell wall. Our data supported that the anchoring of the Cwp84 47-kDa form is presumably due to a reassociation of the secreted protein. Moreover, we showed that the complete maturation of the recombinant protein Cwp84(30-803) is a sequential process beginning at the C-terminal end, followed by one or more cleavages at the N-terminal end. The processing sites of recombinant Cwp84 are likely to be residues Ser-92 and Lys-518. No proteolytic activity was detected with the mature recombinant protease Cwp84(92-518) (47 kDa). In contrast, a fragment including the propeptide (Cwp84(30-518)) displayed proteolytic activity on azocasein and fibronectin. These results showed that Cwp84 is processed essentially at the bacterial cell surface and that its different forms may display different proteolytic activities.

UI MeSH Term Description Entries
D008856 Microscopy, Fluorescence Microscopy of specimens stained with fluorescent dye (usually fluorescein isothiocyanate) or of naturally fluorescent materials, which emit light when exposed to ultraviolet or blue light. Immunofluorescence microscopy utilizes antibodies that are labeled with fluorescent dye. Fluorescence Microscopy,Immunofluorescence Microscopy,Microscopy, Immunofluorescence,Fluorescence Microscopies,Immunofluorescence Microscopies,Microscopies, Fluorescence,Microscopies, Immunofluorescence
D011994 Recombinant Proteins Proteins prepared by recombinant DNA technology. Biosynthetic Protein,Biosynthetic Proteins,DNA Recombinant Proteins,Recombinant Protein,Proteins, Biosynthetic,Proteins, Recombinant DNA,DNA Proteins, Recombinant,Protein, Biosynthetic,Protein, Recombinant,Proteins, DNA Recombinant,Proteins, Recombinant,Recombinant DNA Proteins,Recombinant Proteins, DNA
D003546 Cysteine Endopeptidases ENDOPEPTIDASES which have a cysteine involved in the catalytic process. This group of enzymes is inactivated by CYSTEINE PROTEINASE INHIBITORS such as CYSTATINS and SULFHYDRYL REAGENTS.
D004591 Electrophoresis, Polyacrylamide Gel Electrophoresis in which a polyacrylamide gel is used as the diffusion medium. Polyacrylamide Gel Electrophoresis,SDS-PAGE,Sodium Dodecyl Sulfate-PAGE,Gel Electrophoresis, Polyacrylamide,SDS PAGE,Sodium Dodecyl Sulfate PAGE,Sodium Dodecyl Sulfate-PAGEs
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D015151 Immunoblotting Immunologic method used for detecting or quantifying immunoreactive substances. The substance is identified by first immobilizing it by blotting onto a membrane and then tagging it with labeled antibodies. Dot Immunoblotting,Electroimmunoblotting,Immunoelectroblotting,Reverse Immunoblotting,Immunoblotting, Dot,Immunoblotting, Reverse,Dot Immunoblottings,Electroimmunoblottings,Immunoblottings,Immunoblottings, Dot,Immunoblottings, Reverse,Immunoelectroblottings,Reverse Immunoblottings
D016360 Clostridioides difficile A common inhabitant of the colon flora in human infants and sometimes in adults. The type species Clostridioides difficile is formerly known as Clostridium difficile. It is a causative agent for CLOSTRIDIOIDES INFECTIONS and is associated with PSEUDOMEMBRANOUS ENTEROCOLITIS in patients receiving antibiotic therapy. Clostridium difficile
D051379 Mice The common name for the genus Mus. Mice, House,Mus,Mus musculus,Mice, Laboratory,Mouse,Mouse, House,Mouse, Laboratory,Mouse, Swiss,Mus domesticus,Mus musculus domesticus,Swiss Mice,House Mice,House Mouse,Laboratory Mice,Laboratory Mouse,Mice, Swiss,Swiss Mouse,domesticus, Mus musculus
D053719 Tandem Mass Spectrometry A mass spectrometry technique using two (MS/MS) or more mass analyzers. With two in tandem, the precursor ions are mass-selected by a first mass analyzer, and focused into a collision region where they are then fragmented into product ions which are then characterized by a second mass analyzer. A variety of techniques are used to separate the compounds, ionize them, and introduce them to the first mass analyzer. For example, for in GC-MS/MS, GAS CHROMATOGRAPHY-MASS SPECTROMETRY is involved in separating relatively small compounds by GAS CHROMATOGRAPHY prior to injecting them into an ionization chamber for the mass selection. Mass Spectrometry-Mass Spectrometry,Mass Spectrometry Mass Spectrometry,Mass Spectrometry, Tandem
D019032 Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization A mass spectrometric technique that is used for the analysis of large biomolecules. Analyte molecules are embedded in an excess matrix of small organic molecules that show a high resonant absorption at the laser wavelength used. The matrix absorbs the laser energy, thus inducing a soft disintegration of the sample-matrix mixture into free (gas phase) matrix and analyte molecules and molecular ions. In general, only molecular ions of the analyte molecules are produced, and almost no fragmentation occurs. This makes the method well suited for molecular weight determinations and mixture analysis. Laser Desorption-Ionization Mass Spectrometry, Matrix-Assisted,MALD-MS,MALDI,Mass Spectrometry, Matrix-Assisted Laser Desorption-Ionization,Mass Spectroscopy, Matrix-Assisted Laser Desorption-Ionization,Matrix-Assisted Laser Desorption-Ionization Mass Spectrometry,Spectroscopy, Mass, Matrix-Assisted Laser Desorption-Ionization,MALDI-MS,MS-MALD,SELDI-TOF-MS,Surface Enhanced Laser Desorption Ionization Mass Spectrometry,Laser Desorption Ionization Mass Spectrometry, Matrix Assisted,MALDI MS,Mass Spectrometry, Matrix Assisted Laser Desorption Ionization,Mass Spectroscopy, Matrix Assisted Laser Desorption Ionization,Matrix Assisted Laser Desorption Ionization Mass Spectrometry

Related Publications

Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
March 2015, Acta crystallographica. Section F, Structural biology communications,
Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
December 2009, The Journal of biological chemistry,
Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
February 2013, Anaerobe,
Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
July 2014, Acta crystallographica. Section D, Biological crystallography,
Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
January 2015, PloS one,
Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
October 2007, Journal of bacteriology,
Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
October 2011, FEMS immunology and medical microbiology,
Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
July 2011, Journal of bacteriology,
Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
November 2011, European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik e.V,
Diana ChapetónMontes, and Thomas Candela, and Anne Collignon, and Claire Janoir
June 2020, Molecular and cellular endocrinology,
Copied contents to your clipboard!