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Conditional Protein Splicing Switch in Hyperthermophiles through an Intein-Extein Partnership
Inteins are intervening proteins that undergo an autocatalytic splicing reaction that ligates flanking host protein sequences termed exteins. Some intein-containing proteins have evolved to couple splicing to environmental signals; this represents a new form of posttranslational regulation. Of parti...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Society for Microbiology
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5790916/ https://www.ncbi.nlm.nih.gov/pubmed/29382734 http://dx.doi.org/10.1128/mBio.02304-17 |
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author | Lennon, Christopher W. Stanger, Matthew Banavali, Nilesh K. Belfort, Marlene |
author_facet | Lennon, Christopher W. Stanger, Matthew Banavali, Nilesh K. Belfort, Marlene |
author_sort | Lennon, Christopher W. |
collection | PubMed |
description | Inteins are intervening proteins that undergo an autocatalytic splicing reaction that ligates flanking host protein sequences termed exteins. Some intein-containing proteins have evolved to couple splicing to environmental signals; this represents a new form of posttranslational regulation. Of particular interest is RadA from the archaeon Pyrococcus horikoshii, for which long-range intein-extein interactions block splicing, requiring temperature and single-stranded DNA (ssDNA) substrate to splice rapidly and accurately. Here, we report that splicing of the intein-containing RadA from another archaeon, Thermococcus sibericus, is activated by significantly lower temperatures than is P. horikoshii RadA, consistent with differences in their growth environments. Investigation into variations between T. sibericus and P. horikoshii RadA inteins led to the discovery that a nonconserved region (NCR) of the intein, a flexible loop where a homing endonuclease previously resided, is critical to splicing. Deletion of the NCR leads to a substantial loss in the rate and accuracy of P. horikoshii RadA splicing only within native exteins. The influence of the NCR deletion can be largely overcome by ssDNA, demonstrating that the splicing-competent conformation can be achieved. We present a model whereby the NCR is a flexible hinge which acts as a switch by controlling distant intein-extein interactions that inhibit active site assembly. These results speak to the repurposing of the vestigial endonuclease loop to control an intein-extein partnership, which ultimately allows exquisite adaptation of protein splicing upon changes in the environment. |
format | Online Article Text |
id | pubmed-5790916 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | American Society for Microbiology |
record_format | MEDLINE/PubMed |
spelling | pubmed-57909162018-02-05 Conditional Protein Splicing Switch in Hyperthermophiles through an Intein-Extein Partnership Lennon, Christopher W. Stanger, Matthew Banavali, Nilesh K. Belfort, Marlene mBio Research Article Inteins are intervening proteins that undergo an autocatalytic splicing reaction that ligates flanking host protein sequences termed exteins. Some intein-containing proteins have evolved to couple splicing to environmental signals; this represents a new form of posttranslational regulation. Of particular interest is RadA from the archaeon Pyrococcus horikoshii, for which long-range intein-extein interactions block splicing, requiring temperature and single-stranded DNA (ssDNA) substrate to splice rapidly and accurately. Here, we report that splicing of the intein-containing RadA from another archaeon, Thermococcus sibericus, is activated by significantly lower temperatures than is P. horikoshii RadA, consistent with differences in their growth environments. Investigation into variations between T. sibericus and P. horikoshii RadA inteins led to the discovery that a nonconserved region (NCR) of the intein, a flexible loop where a homing endonuclease previously resided, is critical to splicing. Deletion of the NCR leads to a substantial loss in the rate and accuracy of P. horikoshii RadA splicing only within native exteins. The influence of the NCR deletion can be largely overcome by ssDNA, demonstrating that the splicing-competent conformation can be achieved. We present a model whereby the NCR is a flexible hinge which acts as a switch by controlling distant intein-extein interactions that inhibit active site assembly. These results speak to the repurposing of the vestigial endonuclease loop to control an intein-extein partnership, which ultimately allows exquisite adaptation of protein splicing upon changes in the environment. American Society for Microbiology 2018-01-30 /pmc/articles/PMC5790916/ /pubmed/29382734 http://dx.doi.org/10.1128/mBio.02304-17 Text en Copyright © 2018 Lennon et al. https://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license (https://creativecommons.org/licenses/by/4.0/) . |
spellingShingle | Research Article Lennon, Christopher W. Stanger, Matthew Banavali, Nilesh K. Belfort, Marlene Conditional Protein Splicing Switch in Hyperthermophiles through an Intein-Extein Partnership |
title | Conditional Protein Splicing Switch in Hyperthermophiles through an Intein-Extein Partnership |
title_full | Conditional Protein Splicing Switch in Hyperthermophiles through an Intein-Extein Partnership |
title_fullStr | Conditional Protein Splicing Switch in Hyperthermophiles through an Intein-Extein Partnership |
title_full_unstemmed | Conditional Protein Splicing Switch in Hyperthermophiles through an Intein-Extein Partnership |
title_short | Conditional Protein Splicing Switch in Hyperthermophiles through an Intein-Extein Partnership |
title_sort | conditional protein splicing switch in hyperthermophiles through an intein-extein partnership |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5790916/ https://www.ncbi.nlm.nih.gov/pubmed/29382734 http://dx.doi.org/10.1128/mBio.02304-17 |
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