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Quick and Spontaneous Transformation between [3Fe–4S] and [4Fe–4S] Iron–Sulfur Clusters in the tRNA-Thiolation Enzyme TtuA
Iron–sulfur (Fe–S) clusters are essential cofactors for enzyme activity. These Fe–S clusters are present in structurally diverse forms, including [4Fe–4S] and [3Fe–4S]. Type-identification of the Fe–S cluster is indispensable in understanding the catalytic mechanism of enzymes. However, identifying...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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MDPI
2023
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9821441/ https://www.ncbi.nlm.nih.gov/pubmed/36614280 http://dx.doi.org/10.3390/ijms24010833 |
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author | Ishizaka, Masato Chen, Minghao Narai, Shun Tanaka, Yoshikazu Ose, Toyoyuki Horitani, Masaki Yao, Min |
author_facet | Ishizaka, Masato Chen, Minghao Narai, Shun Tanaka, Yoshikazu Ose, Toyoyuki Horitani, Masaki Yao, Min |
author_sort | Ishizaka, Masato |
collection | PubMed |
description | Iron–sulfur (Fe–S) clusters are essential cofactors for enzyme activity. These Fe–S clusters are present in structurally diverse forms, including [4Fe–4S] and [3Fe–4S]. Type-identification of the Fe–S cluster is indispensable in understanding the catalytic mechanism of enzymes. However, identifying [4Fe–4S] and [3Fe–4S] clusters in particular is challenging because of their rapid transformation in response to oxidation–reduction events. In this study, we focused on the relationship between the Fe–S cluster type and the catalytic activity of a tRNA-thiolation enzyme (TtuA). We reconstituted [4Fe–4S]-TtuA, prepared [3Fe–4S]-TtuA by oxidizing [4Fe–4S]-TtuA under strictly anaerobic conditions, and then observed changes in the Fe–S clusters in the samples and the enzymatic activity in the time-course experiments. Electron paramagnetic resonance analysis revealed that [3Fe–4S]-TtuA spontaneously transforms into [4Fe–4S]-TtuA in minutes to one hour without an additional free Fe source in the solution. Although the TtuA immediately after oxidation of [4Fe–4S]-TtuA was inactive [3Fe–4S]-TtuA, its activity recovered to a significant level compared to [4Fe–4S]-TtuA after one hour, corresponding to an increase of [4Fe–4S]-TtuA in the solution. Our findings reveal that [3Fe–4S]-TtuA is highly inactive and unstable. Moreover, time-course analysis of structural changes and activity under strictly anaerobic conditions further unraveled the Fe–S cluster type used by the tRNA-thiolation enzyme. |
format | Online Article Text |
id | pubmed-9821441 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-98214412023-01-07 Quick and Spontaneous Transformation between [3Fe–4S] and [4Fe–4S] Iron–Sulfur Clusters in the tRNA-Thiolation Enzyme TtuA Ishizaka, Masato Chen, Minghao Narai, Shun Tanaka, Yoshikazu Ose, Toyoyuki Horitani, Masaki Yao, Min Int J Mol Sci Article Iron–sulfur (Fe–S) clusters are essential cofactors for enzyme activity. These Fe–S clusters are present in structurally diverse forms, including [4Fe–4S] and [3Fe–4S]. Type-identification of the Fe–S cluster is indispensable in understanding the catalytic mechanism of enzymes. However, identifying [4Fe–4S] and [3Fe–4S] clusters in particular is challenging because of their rapid transformation in response to oxidation–reduction events. In this study, we focused on the relationship between the Fe–S cluster type and the catalytic activity of a tRNA-thiolation enzyme (TtuA). We reconstituted [4Fe–4S]-TtuA, prepared [3Fe–4S]-TtuA by oxidizing [4Fe–4S]-TtuA under strictly anaerobic conditions, and then observed changes in the Fe–S clusters in the samples and the enzymatic activity in the time-course experiments. Electron paramagnetic resonance analysis revealed that [3Fe–4S]-TtuA spontaneously transforms into [4Fe–4S]-TtuA in minutes to one hour without an additional free Fe source in the solution. Although the TtuA immediately after oxidation of [4Fe–4S]-TtuA was inactive [3Fe–4S]-TtuA, its activity recovered to a significant level compared to [4Fe–4S]-TtuA after one hour, corresponding to an increase of [4Fe–4S]-TtuA in the solution. Our findings reveal that [3Fe–4S]-TtuA is highly inactive and unstable. Moreover, time-course analysis of structural changes and activity under strictly anaerobic conditions further unraveled the Fe–S cluster type used by the tRNA-thiolation enzyme. MDPI 2023-01-03 /pmc/articles/PMC9821441/ /pubmed/36614280 http://dx.doi.org/10.3390/ijms24010833 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Ishizaka, Masato Chen, Minghao Narai, Shun Tanaka, Yoshikazu Ose, Toyoyuki Horitani, Masaki Yao, Min Quick and Spontaneous Transformation between [3Fe–4S] and [4Fe–4S] Iron–Sulfur Clusters in the tRNA-Thiolation Enzyme TtuA |
title | Quick and Spontaneous Transformation between [3Fe–4S] and [4Fe–4S] Iron–Sulfur Clusters in the tRNA-Thiolation Enzyme TtuA |
title_full | Quick and Spontaneous Transformation between [3Fe–4S] and [4Fe–4S] Iron–Sulfur Clusters in the tRNA-Thiolation Enzyme TtuA |
title_fullStr | Quick and Spontaneous Transformation between [3Fe–4S] and [4Fe–4S] Iron–Sulfur Clusters in the tRNA-Thiolation Enzyme TtuA |
title_full_unstemmed | Quick and Spontaneous Transformation between [3Fe–4S] and [4Fe–4S] Iron–Sulfur Clusters in the tRNA-Thiolation Enzyme TtuA |
title_short | Quick and Spontaneous Transformation between [3Fe–4S] and [4Fe–4S] Iron–Sulfur Clusters in the tRNA-Thiolation Enzyme TtuA |
title_sort | quick and spontaneous transformation between [3fe–4s] and [4fe–4s] iron–sulfur clusters in the trna-thiolation enzyme ttua |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9821441/ https://www.ncbi.nlm.nih.gov/pubmed/36614280 http://dx.doi.org/10.3390/ijms24010833 |
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