Structural and Biochemical Characterization of Thioredoxin-2 from <i>Deinococcus radiodurans</i>

Thioredoxin (Trx), a ubiquitous protein showing disulfide reductase activity, plays critical roles in cellular redox control and oxidative stress response. Trx is a member of the Trx system, comprising Trx, Trx reductase (TrxR), and a cognate reductant (generally reduced nicotinamide adenine dinucle...

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Autores principales:	Min-Kyu Kim, Lei Zhao, Soyoung Jeong, Jing Zhang, Jong-Hyun Jung, Ho Seong Seo, Jong-il Choi, Sangyong Lim
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	thioredoxin Trx2 <i>D. radiodurans</i> crystal structure disulfide reduction Therapeutics. Pharmacology RM1-950
Acceso en línea:	https://doaj.org/article/b312dcff79ea479791b3360d711de152
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spelling	oai:doaj.org-article:b312dcff79ea479791b3360d711de1522021-11-25T16:29:44ZStructural and Biochemical Characterization of Thioredoxin-2 from <i>Deinococcus radiodurans</i>10.3390/antiox101118432076-3921https://doaj.org/article/b312dcff79ea479791b3360d711de1522021-11-01T00:00:00Zhttps://www.mdpi.com/2076-3921/10/11/1843https://doaj.org/toc/2076-3921Thioredoxin (Trx), a ubiquitous protein showing disulfide reductase activity, plays critical roles in cellular redox control and oxidative stress response. Trx is a member of the Trx system, comprising Trx, Trx reductase (TrxR), and a cognate reductant (generally reduced nicotinamide adenine dinucleotide phosphate, NADPH). Bacterial Trx1 contains only the Trx-fold domain, in which the active site CXXC motif that is critical for the disulfide reduction activity is located. Bacterial Trx2 contains an N-terminal extension, which forms a zinc-finger domain, including two additional CXXC motifs. The multi-stress resistant bacterium <i>Deinococcus radiodurans</i> encodes both Trx1 (DrTrx1) and Trx2 (DrTrx2), which act as members of the enzymatic antioxidant systems. In this study, we constructed Δ<i>drtrx1</i> and Δ<i>drtrx2</i> mutants and examined their survival rates under H<sub>2</sub>O<sub>2</sub> treated conditions. Both <i>drtrx1</i> and <i>drtrx2</i> genes were induced following H<sub>2</sub>O<sub>2</sub> treatment, and the Δ<i>drtrx1</i> and Δ<i>drtrx2</i> mutants showed a decrease in resistance toward H<sub>2</sub>O<sub>2</sub>, compared to the wild-type. Native DrTrx1 and DrTrx2 clearly displayed insulin and DTNB reduction activity, whereas mutant DrTrx1 and DrTrx2, which harbors the substitution of conserved cysteine to serine in its active site CXXC motif, showed almost no reduction activity. Mutations in the zinc binding cysteines did not fully eliminate the reduction activities of DrTrx2. Furthermore, we solved the crystal structure of full-length DrTrx2 at 1.96 Å resolution. The N-terminal zinc-finger domain of Trx2 is thought to be involved in Trx-target interaction and, from our DrTrx2 structure, the orientation of the zinc-finger domain of DrTrx2 and its interdomain interaction, between the Trx-fold domain and the zinc-finger domain, is clearly distinguished from those of the other Trx2 structures.Min-Kyu KimLei ZhaoSoyoung JeongJing ZhangJong-Hyun JungHo Seong SeoJong-il ChoiSangyong LimMDPI AGarticlethioredoxinTrx2<i>D. radiodurans</i>crystal structuredisulfide reductionTherapeutics. PharmacologyRM1-950ENAntioxidants, Vol 10, Iss 1843, p 1843 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	thioredoxin Trx2 <i>D. radiodurans</i> crystal structure disulfide reduction Therapeutics. Pharmacology RM1-950
spellingShingle	thioredoxin Trx2 <i>D. radiodurans</i> crystal structure disulfide reduction Therapeutics. Pharmacology RM1-950 Min-Kyu Kim Lei Zhao Soyoung Jeong Jing Zhang Jong-Hyun Jung Ho Seong Seo Jong-il Choi Sangyong Lim Structural and Biochemical Characterization of Thioredoxin-2 from <i>Deinococcus radiodurans</i>
description	Thioredoxin (Trx), a ubiquitous protein showing disulfide reductase activity, plays critical roles in cellular redox control and oxidative stress response. Trx is a member of the Trx system, comprising Trx, Trx reductase (TrxR), and a cognate reductant (generally reduced nicotinamide adenine dinucleotide phosphate, NADPH). Bacterial Trx1 contains only the Trx-fold domain, in which the active site CXXC motif that is critical for the disulfide reduction activity is located. Bacterial Trx2 contains an N-terminal extension, which forms a zinc-finger domain, including two additional CXXC motifs. The multi-stress resistant bacterium <i>Deinococcus radiodurans</i> encodes both Trx1 (DrTrx1) and Trx2 (DrTrx2), which act as members of the enzymatic antioxidant systems. In this study, we constructed Δ<i>drtrx1</i> and Δ<i>drtrx2</i> mutants and examined their survival rates under H<sub>2</sub>O<sub>2</sub> treated conditions. Both <i>drtrx1</i> and <i>drtrx2</i> genes were induced following H<sub>2</sub>O<sub>2</sub> treatment, and the Δ<i>drtrx1</i> and Δ<i>drtrx2</i> mutants showed a decrease in resistance toward H<sub>2</sub>O<sub>2</sub>, compared to the wild-type. Native DrTrx1 and DrTrx2 clearly displayed insulin and DTNB reduction activity, whereas mutant DrTrx1 and DrTrx2, which harbors the substitution of conserved cysteine to serine in its active site CXXC motif, showed almost no reduction activity. Mutations in the zinc binding cysteines did not fully eliminate the reduction activities of DrTrx2. Furthermore, we solved the crystal structure of full-length DrTrx2 at 1.96 Å resolution. The N-terminal zinc-finger domain of Trx2 is thought to be involved in Trx-target interaction and, from our DrTrx2 structure, the orientation of the zinc-finger domain of DrTrx2 and its interdomain interaction, between the Trx-fold domain and the zinc-finger domain, is clearly distinguished from those of the other Trx2 structures.
format	article
author	Min-Kyu Kim Lei Zhao Soyoung Jeong Jing Zhang Jong-Hyun Jung Ho Seong Seo Jong-il Choi Sangyong Lim
author_facet	Min-Kyu Kim Lei Zhao Soyoung Jeong Jing Zhang Jong-Hyun Jung Ho Seong Seo Jong-il Choi Sangyong Lim
author_sort	Min-Kyu Kim
title	Structural and Biochemical Characterization of Thioredoxin-2 from <i>Deinococcus radiodurans</i>
title_short	Structural and Biochemical Characterization of Thioredoxin-2 from <i>Deinococcus radiodurans</i>
title_full	Structural and Biochemical Characterization of Thioredoxin-2 from <i>Deinococcus radiodurans</i>
title_fullStr	Structural and Biochemical Characterization of Thioredoxin-2 from <i>Deinococcus radiodurans</i>
title_full_unstemmed	Structural and Biochemical Characterization of Thioredoxin-2 from <i>Deinococcus radiodurans</i>
title_sort	structural and biochemical characterization of thioredoxin-2 from <i>deinococcus radiodurans</i>
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/b312dcff79ea479791b3360d711de152
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Structural and Biochemical Characterization of Thioredoxin-2 from <i>Deinococcus radiodurans</i>

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