1H, 13C and 15N resonance assignment of the mature form of monothiol glutaredoxin 1 from the pathogen Trypanosoma brucei

Sturlese, Mattia; Lelli, Moreno; Manta, Bruno; Mammi, Stefano; Comini, Marcelo A.; Bellanda, Massimo

doi:10.1007/s12104-014-9561-3

Cited by 5 publications

(10 citation statements)

References 12 publications

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“…The N-terminal half (G48 to P59) displayed low propensity to adopt a regular secondary structure whereas the C-terminal half (P63 to V75) was predicted to form regular secondary structures 8 . The disordered nature of the N-terminal extension was suggested previously 8 , 23 and is conclusively shown here by several new pieces of evidence obtained by NMR. The residues belonging to the NTE show a poor dispersion of the proton chemical shifts in the 1 H- 15 N HSQC spectrum.…”

Section: Resultssupporting

confidence: 84%

“…All recombinant proteins were produced as fusion proteins to E . coli Trx and purified according to the procedure described in 23 . Briefly, transformed E .…”

Section: Methodsmentioning

confidence: 99%

“…The preparation of FL 1CGrx1 for NMR analysis and the respective resonance assignment was previously reported 23 . Briefly, the samples for NMR experiments were prepared in 50 mM sodium phosphate, pH 7.0, 150 mM NaCl, and 10 mM DTT.…”

Section: Methodsmentioning

confidence: 99%

“…The resonance assignment of FL 1CGrx1 was previously reported 23 and its structure was solved following the same procedure used to solve the structure of the truncated Δ76 version 8 . Briefly, backbone dihedral angle constraints were derived from 15 N, 13 C′, 13 Cα, 13 Cβ, and Hα chemical shifts using TALOS+ 35 .…”

Section: Methodsmentioning

confidence: 99%

“…More recently we reported the NMR resonance assignment of FL 1CGrx1. The poor dispersion of the peaks in the proton dimension of the HSQC spectrum for residues belonging to this tail, suggested a non-globular nature of the N-terminal extension of the protein 23 . The presence of a long, disordered extension before or after the conserved Grx domain is not a common feature in Grx.…”

Section: Introductionmentioning

confidence: 99%

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The lineage-specific, intrinsically disordered N-terminal extension of monothiol glutaredoxin 1 from trypanosomes contains a regulatory region

Sturlese

Manta

Bertarello

et al. 2018

Sci Rep

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Glutaredoxins (Grx) are small proteins conserved throughout all the kingdoms of life that are engaged in a wide variety of biological processes and share a common thioredoxin-fold. Among them, class II Grx are redox-inactive proteins involved in iron-sulfur (FeS) metabolism. They contain a single thiol group in their active site and use low molecular mass thiols such as glutathione as ligand for binding FeS-clusters. In this study, we investigated molecular aspects of 1CGrx1 from the pathogenic parasite Trypanosoma brucei brucei, a mitochondrial class II Grx that fulfills an indispensable role in vivo. Mitochondrial 1CGrx1 from trypanosomes differs from orthologues in several features including the presence of a parasite-specific N-terminal extension (NTE) whose role has yet to be elucidated. Previously we have solved the structure of a truncated form of 1CGrx1 containing only the conserved glutaredoxin domain but lacking the NTE. Our aim here is to investigate the effect of the NTE on the conformation of the protein. We therefore solved the NMR structure of the full-length protein, which reveals subtle but significant differences with the structure of the NTE-less form. By means of different experimental approaches, the NTE proved to be intrinsically disordered and not involved in the non-redox dependent protein dimerization, as previously suggested. Interestingly, the portion comprising residues 65–76 of the NTE modulates the conformational dynamics of the glutathione-binding pocket, which may play a role in iron-sulfur cluster assembly and delivery. Furthermore, we disclosed that the class II-strictly conserved loop that precedes the active site is critical for stabilizing the protein structure. So far, this represents the first communication of a Grx containing an intrinsically disordered region that defines a new protein subgroup within class II Grx.

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Section: Resultssupporting

confidence: 84%

“…All recombinant proteins were produced as fusion proteins to E . coli Trx and purified according to the procedure described in 23 . Briefly, transformed E .…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

The lineage-specific, intrinsically disordered N-terminal extension of monothiol glutaredoxin 1 from trypanosomes contains a regulatory region

Sturlese

Manta

Bertarello

et al. 2018

Sci Rep

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1H, 13C and 15N resonance assignment of the cytosolic dithiol glutaredoxin 1 from the pathogen Trypanosoma brucei

et al. 2015

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Trypanosomatids are parasites responsible for several tropical and subtropical diseases, such as Chaga's disease, sleeping sickness and Leishmaniasis. In contrast to the mammalian host, the thiol-redox metabolism of these pathogens depends on trypanothione [bis-glutathionylspermidine, T(SH)2] instead of glutathione (GSH) providing a set of lineage-specific proteins as drug target candidates. Glutaredoxins (Grx) are ubiquitous small thiol-disulfide oxidoreductases that belong to the thioredoxin-fold family. They play a central role in redox homeostasis and iron sulfur-cluster biogenesis. Each species, including trypanosomes, possesses its own set of isoforms distributed in different subcellular compartments. The genome of trypanosomatids encodes for two class I (dithiolic) Grxs named 2-C-Grx1 and 2-C-Grx2. Both proteins were shown to efficiently reduce different disulfides at the expenses of T(SH)2 using a mechanism that involves the two cysteines in the active site. Moreover, the cytosolic Trypanosoma brucei 2-C-Grx1 but not the mitochondrial 2-C-Grx2 was able to coordinate an iron-sulfur cluster with T(SH)2 or GSH as ligand. As a first step to unravel the structural basis for the specificity observed in the trypanosomal glutaredoxins, we present here the NMR resonance assignment of 2-C-Grx1 from the parasite T. brucei brucei.

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Intrinsically Disordered Region Modulates Ligand Binding in Glutaredoxin 1 from Trypanosoma Brucei

et al. 2021

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Glutaredoxins are small proteins that share a common well-conserved thioredoxin-fold and participate in a wide variety of biological processes. Among them, class II Grx are redox-inactive proteins involved in iron−sulfur (Fe-S) metabolism. In the present work, we report different structural and dynamics aspects of 1CGrx1 from the pathogenic parasite Trypanosoma brucei that differentiate it from other orthologues by the presence of a parasite-specific unstructured N-terminal extension whose role has not been fully elucidated yet. Previous nuclear magnetic resonance (NMR) studies revealed significant differences with respect to the mutant lacking the disordered tail. Herein, we have performed atomistic molecular dynamics simulations that, complementary to NMR studies, confirm the intrinsically disordered nature of the N-terminal extension. Moreover, we confirm the main role of these residues in modulating the conformational dynamics of the glutathione-binding pocket. We observe that the N-terminal extension modifies the ligand cavity stiffening it by specific interactions that ultimately modulate its intrinsic flexibility, which may modify its role in the storage and/or transfer of preformed iron−sulfur clusters. These unique structural and dynamics aspects of Trypanosoma brucei 1CGrx1 differentiate it from other orthologues and could have functional relevance. In this way, our results encourage the study of other similar protein folding families with intrinsically disordered regions whose functional roles are still unrevealed and the screening of potential 1CGrx1 inhibitors as antitrypanosomal drug candidates.

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1H, 13C and 15N resonance assignment of the mature form of monothiol glutaredoxin 1 from the pathogen Trypanosoma brucei

Cited by 5 publications

References 12 publications

The lineage-specific, intrinsically disordered N-terminal extension of monothiol glutaredoxin 1 from trypanosomes contains a regulatory region

The lineage-specific, intrinsically disordered N-terminal extension of monothiol glutaredoxin 1 from trypanosomes contains a regulatory region

1H, 13C and 15N resonance assignment of the cytosolic dithiol glutaredoxin 1 from the pathogen Trypanosoma brucei

Intrinsically Disordered Region Modulates Ligand Binding in Glutaredoxin 1 from Trypanosoma Brucei

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