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In vivo probing of SECIS-dependent selenocysteine translation in Archaea

Cotranslational insertion of selenocysteine (Sec) proceeds by recoding UGA to a sense codon. This recoding is governed by the Sec insertion sequence (SECIS) element, an RNA structure on the mRNA, but size, location, structure determinants, and mechanism differ for Bacteria, Eukarya, and Archaea. For...

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Autores principales: Peiter, Nils, Rother, Michael
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Life Science Alliance LLC 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9622424/
https://www.ncbi.nlm.nih.gov/pubmed/36316034
http://dx.doi.org/10.26508/lsa.202201676
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author Peiter, Nils
Rother, Michael
author_facet Peiter, Nils
Rother, Michael
author_sort Peiter, Nils
collection PubMed
description Cotranslational insertion of selenocysteine (Sec) proceeds by recoding UGA to a sense codon. This recoding is governed by the Sec insertion sequence (SECIS) element, an RNA structure on the mRNA, but size, location, structure determinants, and mechanism differ for Bacteria, Eukarya, and Archaea. For Archaea, the structure–function relation of the SECIS is poorly understood, as only rather laborious experimental approaches are established. Furthermore, these methods do not allow for quantitative probing of Sec insertion. In order to overcome these limitations, we engineered bacterial β-lactamase into an archaeal selenoprotein, thereby establishing a reporter system, which correlates enzyme activity to Sec insertion. Using this system, in vivo Sec insertion depending on the availability of selenium and the presence of a SECIS element was assessed in Methanococcus maripaludis. Furthermore, a minimal SECIS element required for Sec insertion in M. maripaludis was defined and a conserved structural motif shown to be essential for function. Besides developing a convenient tool for selenium research, converting a bacterial enzyme into an archaeal selenoprotein provides proof of concept that novel selenoproteins can be engineered in Archaea.
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spelling pubmed-96224242022-11-02 In vivo probing of SECIS-dependent selenocysteine translation in Archaea Peiter, Nils Rother, Michael Life Sci Alliance Research Articles Cotranslational insertion of selenocysteine (Sec) proceeds by recoding UGA to a sense codon. This recoding is governed by the Sec insertion sequence (SECIS) element, an RNA structure on the mRNA, but size, location, structure determinants, and mechanism differ for Bacteria, Eukarya, and Archaea. For Archaea, the structure–function relation of the SECIS is poorly understood, as only rather laborious experimental approaches are established. Furthermore, these methods do not allow for quantitative probing of Sec insertion. In order to overcome these limitations, we engineered bacterial β-lactamase into an archaeal selenoprotein, thereby establishing a reporter system, which correlates enzyme activity to Sec insertion. Using this system, in vivo Sec insertion depending on the availability of selenium and the presence of a SECIS element was assessed in Methanococcus maripaludis. Furthermore, a minimal SECIS element required for Sec insertion in M. maripaludis was defined and a conserved structural motif shown to be essential for function. Besides developing a convenient tool for selenium research, converting a bacterial enzyme into an archaeal selenoprotein provides proof of concept that novel selenoproteins can be engineered in Archaea. Life Science Alliance LLC 2022-10-31 /pmc/articles/PMC9622424/ /pubmed/36316034 http://dx.doi.org/10.26508/lsa.202201676 Text en © 2022 Peiter and Rother https://creativecommons.org/licenses/by/4.0/This article is available under a Creative Commons License (Attribution 4.0 International, as described at https://creativecommons.org/licenses/by/4.0/).
spellingShingle Research Articles
Peiter, Nils
Rother, Michael
In vivo probing of SECIS-dependent selenocysteine translation in Archaea
title In vivo probing of SECIS-dependent selenocysteine translation in Archaea
title_full In vivo probing of SECIS-dependent selenocysteine translation in Archaea
title_fullStr In vivo probing of SECIS-dependent selenocysteine translation in Archaea
title_full_unstemmed In vivo probing of SECIS-dependent selenocysteine translation in Archaea
title_short In vivo probing of SECIS-dependent selenocysteine translation in Archaea
title_sort in vivo probing of secis-dependent selenocysteine translation in archaea
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9622424/
https://www.ncbi.nlm.nih.gov/pubmed/36316034
http://dx.doi.org/10.26508/lsa.202201676
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