A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein.
Colicins are protein antibiotics synthesised by Escherichia coli strains to target and kill related bacteria. To prevent host suicide, colicins are inactivated by binding to immunity proteins. Despite their high avidity (K(d) ≈ fM, lifetime ≈ 4 days), immunity protein release is a pre-requisite of c...
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Public Library of Science (PLoS)
2013-01-01
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| Series: | PLoS Biology |
| Online Access: | https://doi.org/10.1371/journal.pbio.1001489 |
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| author | Oliver E Farrance Eleanore Hann Renata Kaminska Nicholas G Housden Sasha R Derrington Colin Kleanthous Sheena E Radford David J Brockwell |
| author_facet | Oliver E Farrance Eleanore Hann Renata Kaminska Nicholas G Housden Sasha R Derrington Colin Kleanthous Sheena E Radford David J Brockwell |
| author_sort | Oliver E Farrance |
| collection | DOAJ |
| description | Colicins are protein antibiotics synthesised by Escherichia coli strains to target and kill related bacteria. To prevent host suicide, colicins are inactivated by binding to immunity proteins. Despite their high avidity (K(d) ≈ fM, lifetime ≈ 4 days), immunity protein release is a pre-requisite of colicin intoxication, which occurs on a timescale of minutes. Here, by measuring the dynamic force spectrum of the dissociation of the DNase domain of colicin E9 (E9) and immunity protein 9 (Im9) complex using an atomic force microscope we show that application of low forces (<20 pN) increases the rate of complex dissociation 10(6)-fold, to a timescale (lifetime ≈ 10 ms) compatible with intoxication. We term this catastrophic force-triggered increase in off-rate a trip bond. Using mutational analysis, we elucidate the mechanism of this switch in affinity. We show that the N-terminal region of E9, which has sparse contacts with the hydrophobic core, is linked to an allosteric activator region in E9 (residues 21-30) whose remodelling triggers immunity protein release. Diversion of the force transduction pathway by the introduction of appropriately positioned disulfide bridges yields a force resistant complex with a lifetime identical to that measured by ensemble techniques. A trip switch within E9 is ideal for its function as it allows bipartite complex affinity, whereby the stable colicin:immunity protein complex required for host protection can be readily converted to a kinetically unstable complex whose dissociation is necessary for cellular invasion and competitor death. More generally, the observation of two force phenotypes for the E9:Im9 complex demonstrates that force can re-sculpt the underlying energy landscape, providing new opportunities to modulate biological reactions in vivo; this rationalises the commonly observed discrepancy between off-rates measured by dynamic force spectroscopy and ensemble methods. |
| format | Article |
| id | doaj-art-971c0dd9777f4a4593fa9ceffca2a088 |
| institution | OA Journals |
| issn | 1544-9173 1545-7885 |
| language | English |
| publishDate | 2013-01-01 |
| publisher | Public Library of Science (PLoS) |
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| series | PLoS Biology |
| spelling | doaj-art-971c0dd9777f4a4593fa9ceffca2a0882025-08-20T02:22:49ZengPublic Library of Science (PLoS)PLoS Biology1544-91731545-78852013-01-01112e100148910.1371/journal.pbio.1001489A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein.Oliver E FarranceEleanore HannRenata KaminskaNicholas G HousdenSasha R DerringtonColin KleanthousSheena E RadfordDavid J BrockwellColicins are protein antibiotics synthesised by Escherichia coli strains to target and kill related bacteria. To prevent host suicide, colicins are inactivated by binding to immunity proteins. Despite their high avidity (K(d) ≈ fM, lifetime ≈ 4 days), immunity protein release is a pre-requisite of colicin intoxication, which occurs on a timescale of minutes. Here, by measuring the dynamic force spectrum of the dissociation of the DNase domain of colicin E9 (E9) and immunity protein 9 (Im9) complex using an atomic force microscope we show that application of low forces (<20 pN) increases the rate of complex dissociation 10(6)-fold, to a timescale (lifetime ≈ 10 ms) compatible with intoxication. We term this catastrophic force-triggered increase in off-rate a trip bond. Using mutational analysis, we elucidate the mechanism of this switch in affinity. We show that the N-terminal region of E9, which has sparse contacts with the hydrophobic core, is linked to an allosteric activator region in E9 (residues 21-30) whose remodelling triggers immunity protein release. Diversion of the force transduction pathway by the introduction of appropriately positioned disulfide bridges yields a force resistant complex with a lifetime identical to that measured by ensemble techniques. A trip switch within E9 is ideal for its function as it allows bipartite complex affinity, whereby the stable colicin:immunity protein complex required for host protection can be readily converted to a kinetically unstable complex whose dissociation is necessary for cellular invasion and competitor death. More generally, the observation of two force phenotypes for the E9:Im9 complex demonstrates that force can re-sculpt the underlying energy landscape, providing new opportunities to modulate biological reactions in vivo; this rationalises the commonly observed discrepancy between off-rates measured by dynamic force spectroscopy and ensemble methods.https://doi.org/10.1371/journal.pbio.1001489 |
| spellingShingle | Oliver E Farrance Eleanore Hann Renata Kaminska Nicholas G Housden Sasha R Derrington Colin Kleanthous Sheena E Radford David J Brockwell A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein. PLoS Biology |
| title | A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein. |
| title_full | A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein. |
| title_fullStr | A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein. |
| title_full_unstemmed | A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein. |
| title_short | A force-activated trip switch triggers rapid dissociation of a colicin from its immunity protein. |
| title_sort | force activated trip switch triggers rapid dissociation of a colicin from its immunity protein |
| url | https://doi.org/10.1371/journal.pbio.1001489 |
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