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Yorodumi- PDB-4cu5: C-terminal domain of endolysin from phage CD27L is a trigger and ... -
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-Basic information
Entry | Database: PDB / ID: 4cu5 | ||||||
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Title | C-terminal domain of endolysin from phage CD27L is a trigger and release factor | ||||||
Components | ENDOLYSIN | ||||||
Keywords | HYDROLASE / BACTERIAL LYSIS / BACTERIOPHAGE / AUTOPROTEOLYSIS | ||||||
Function / homology | Function and homology information N-acetylmuramoyl-L-alanine amidase activity / peptidoglycan catabolic process / metal ion binding Similarity search - Function | ||||||
Biological species | CLOSTRIDIUM PHAGE PHICD27 (virus) | ||||||
Method | X-RAY DIFFRACTION / SYNCHROTRON / SIRAS / Resolution: 2.24 Å | ||||||
Authors | Dunne, M. / Mertens, H.D.T. / Garefalaki, V. / Jeffries, C.M. / Thompson, A. / Lemke, E.A. / Svergun, D.I. / Mayer, M.J. / Narbad, A. / Meijers, R. | ||||||
Citation | Journal: PLoS Pathog / Year: 2014 Title: The CD27L and CTP1L endolysins targeting Clostridia contain a built-in trigger and release factor. Authors: Matthew Dunne / Haydyn D T Mertens / Vasiliki Garefalaki / Cy M Jeffries / Andrew Thompson / Edward A Lemke / Dmitri I Svergun / Melinda J Mayer / Arjan Narbad / Rob Meijers / Abstract: The bacteriophage ΦCD27 is capable of lysing Clostridium difficile, a pathogenic bacterium that is a major cause for nosocomial infection. A recombinant CD27L endolysin lyses C. difficile in vitro, ...The bacteriophage ΦCD27 is capable of lysing Clostridium difficile, a pathogenic bacterium that is a major cause for nosocomial infection. A recombinant CD27L endolysin lyses C. difficile in vitro, and represents a promising alternative as a bactericide. To better understand the lysis mechanism, we have determined the crystal structure of an autoproteolytic fragment of the CD27L endolysin. The structure covers the C-terminal domain of the endolysin, and represents a novel fold that is identified in a number of lysins that target Clostridia bacteria. The structure indicates endolysin cleavage occurs at the stem of the linker connecting the catalytic domain with the C-terminal domain. We also solved the crystal structure of the C-terminal domain of a slow cleaving mutant of the CTP1L endolysin that targets C. tyrobutyricum. Two distinct dimerization modes are observed in the crystal structures for both endolysins, despite a sequence identity of only 22% between the domains. The dimers are validated to be present for the full length protein in solution by right angle light scattering, small angle X-ray scattering and cross-linking experiments using the cross-linking amino acid p-benzoyl-L-phenylalanine (pBpa). Mutagenesis on residues contributing to the dimer interfaces indicates that there is a link between the dimerization modes and the autocleavage mechanism. We show that for the CTP1L endolysin, there is a reduction in lysis efficiency that is proportional to the cleavage efficiency. We propose a model for endolysin triggering, where the extended dimer presents the inactive state, and a switch to the side-by-side dimer triggers the cleavage of the C-terminal domain. This leads to the release of the catalytic portion of the endolysin, enabling the efficient digestion of the bacterial cell wall. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 4cu5.cif.gz | 117.1 KB | Display | PDBx/mmCIF format |
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PDB format | pdb4cu5.ent.gz | 94.3 KB | Display | PDB format |
PDBx/mmJSON format | 4cu5.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 4cu5_validation.pdf.gz | 447.5 KB | Display | wwPDB validaton report |
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Full document | 4cu5_full_validation.pdf.gz | 450.1 KB | Display | |
Data in XML | 4cu5_validation.xml.gz | 24.1 KB | Display | |
Data in CIF | 4cu5_validation.cif.gz | 35.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/cu/4cu5 ftp://data.pdbj.org/pub/pdb/validation_reports/cu/4cu5 | HTTPS FTP |
-Related structure data
-Links
-Assembly
Deposited unit |
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Unit cell |
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Noncrystallographic symmetry (NCS) | NCS oper:
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-Components
#1: Protein | Mass: 9598.022 Da / Num. of mol.: 6 / Fragment: C-TERMINAL DOMAIN, RESIDUES 186-270 Source method: isolated from a genetically manipulated source Source: (gene. exp.) CLOSTRIDIUM PHAGE PHICD27 (virus) / Production host: ESCHERICHIA COLI (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: B6SBV8 #2: Water | ChemComp-HOH / | |
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-Experimental details
-Experiment
Experiment | Method: X-RAY DIFFRACTION / Number of used crystals: 1 |
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-Sample preparation
Crystal | Density Matthews: 2.3 Å3/Da / Density % sol: 46 % / Description: NONE |
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Crystal grow | pH: 8 / Details: 10 % PEG 20K AND 20 MM TRIS PH 8.0 |
-Data collection
Diffraction | Mean temperature: 100 K |
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Diffraction source | Source: SYNCHROTRON / Site: SOLEIL / Beamline: PROXIMA 1 / Wavelength: 0.97 |
Detector | Type: ADSC QUANTUM 315r / Detector: CCD / Details: KB MIRRORS |
Radiation | Monochromator: SI 1 1 1 / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray |
Radiation wavelength | Wavelength: 0.97 Å / Relative weight: 1 |
Reflection | Resolution: 2.24→30 Å / Num. obs: 24189 / % possible obs: 97.9 % / Observed criterion σ(I): 0 / Redundancy: 2.7 % / Rmerge(I) obs: 0.13 / Net I/σ(I): 7 |
Reflection shell | Resolution: 2.24→2.37 Å / Redundancy: 2.6 % / Rmerge(I) obs: 0.6 / Mean I/σ(I) obs: 2 / % possible all: 93.8 |
-Processing
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Refinement | Method to determine structure: SIRAS Starting model: NONE Resolution: 2.24→33.1 Å / Cor.coef. Fo:Fc: 0.95 / Cor.coef. Fo:Fc free: 0.909 / SU B: 7.392 / SU ML: 0.182 / Cross valid method: THROUGHOUT / ESU R: 0.354 / ESU R Free: 0.242 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS.
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Solvent computation | Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 33.874 Å2
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Refinement step | Cycle: LAST / Resolution: 2.24→33.1 Å
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