The CD27L and CTP1L endolysins targeting Clostridia contain a built-in trigger and release factor.
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...
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Public Library of Science (PLoS)
2014-07-01
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| Series: | PLoS Pathogens |
| Online Access: | https://journals.plos.org/plospathogens/article/file?id=10.1371/journal.ppat.1004228&type=printable |
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| author | 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 |
| author_facet | 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 |
| author_sort | Matthew Dunne |
| collection | DOAJ |
| description | 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. |
| format | Article |
| id | doaj-art-76e1091774d84f3ca598478c1eec7367 |
| institution | Kabale University |
| issn | 1553-7366 1553-7374 |
| language | English |
| publishDate | 2014-07-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS Pathogens |
| spelling | doaj-art-76e1091774d84f3ca598478c1eec73672025-08-20T03:46:43ZengPublic Library of Science (PLoS)PLoS Pathogens1553-73661553-73742014-07-01107e100422810.1371/journal.ppat.1004228The CD27L and CTP1L endolysins targeting Clostridia contain a built-in trigger and release factor.Matthew DunneHaydyn D T MertensVasiliki GarefalakiCy M JeffriesAndrew ThompsonEdward A LemkeDmitri I SvergunMelinda J MayerArjan NarbadRob MeijersThe 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.https://journals.plos.org/plospathogens/article/file?id=10.1371/journal.ppat.1004228&type=printable |
| spellingShingle | 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 The CD27L and CTP1L endolysins targeting Clostridia contain a built-in trigger and release factor. PLoS Pathogens |
| title | The CD27L and CTP1L endolysins targeting Clostridia contain a built-in trigger and release factor. |
| title_full | The CD27L and CTP1L endolysins targeting Clostridia contain a built-in trigger and release factor. |
| title_fullStr | The CD27L and CTP1L endolysins targeting Clostridia contain a built-in trigger and release factor. |
| title_full_unstemmed | The CD27L and CTP1L endolysins targeting Clostridia contain a built-in trigger and release factor. |
| title_short | The CD27L and CTP1L endolysins targeting Clostridia contain a built-in trigger and release factor. |
| title_sort | cd27l and ctp1l endolysins targeting clostridia contain a built in trigger and release factor |
| url | https://journals.plos.org/plospathogens/article/file?id=10.1371/journal.ppat.1004228&type=printable |
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