Investigation of in Vivo Diferric Tyrosyl Radical Formation in Saccharomyces cerevisiae Rnr2 Protein

Zhang, Yan; Liu, Huan; Wu, Xianghua; An, Xiuxiang; Stubbe, JoAnne; Huang, Mingxia

doi:10.1074/jbc.m111.294074

Cited by 49 publications

(89 citation statements)

References 66 publications

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“…An electron transfer process coupled with tightly regulated conformational rearrangements can therefore be proposed to occur in the anamorsin-Ndor1 complex ( Our study lays out the molecular basis for the comprehension of the two functional processes in which the anamorsin-Ndor1 complex is implicated, i.e., assembly of Fe/S proteins and regulation of cell survival/death mechanisms. From our data we can propose that the electron transfer process responsible for the assembly of ISC (13) and diferric (17) proteins occurs within a stable complex without the dissociation of the two protein partners but just through the modulation of the interactions of the domains involved in the electron transfer process. Although the molecular targets of the electron transfer flow generated by this protein-protein complex are not yet defined, suggestions for the targets of the electron flow include the conversion of the sulfur of cysteine (formally S 0 ) to the sulfide (S 2− ) present in Fe/S clusters and/or the reductive coupling of two [2Fe-2S] clusters to form a [4Fe-4S] cluster (13).…”

Section: Discussionmentioning

confidence: 79%

“…This is based on the structural characterization of the FMN-binding domain of Ndor1 and of the C-terminal region (linker and CIAPIN1 domain) of anamorsin containing the [2Fe-2S] cluster in the oxidized state ([2Fe-2S]-anamorsin, hereafter) and on the identification of the protein regions between [2Fe-2S]-anamorsin and the FMN-binding domain of Ndor1 responsible for the formation of their stable complex and of those regions interacting in the electron transfer process. The molecular model of the electron transfer process proposed here provides significant information on the functional processes in which the anamorsinNdor1 interaction has been implicated, i.e., the assembly of ISCs (13) and diferric (17) proteins and the regulation of cell survival/ death mechanisms (15,16). This article is a PNAS Direct Submission.…”

mentioning

confidence: 96%

“…The Tah18-Dre2 interaction is essential for yeast viability and it has been implicated in cellular death regulation mechanisms (16). This complex was proposed to provide electrons necessary for the CIA machinery (13) and for assembling the diferric tyrosyl radical cofactor of ribonucleotide reductase, Rnr2 (17). However, the molecular basis of these processes has not been elucidated.…”

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confidence: 99%

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Molecular view of an electron transfer process essential for iron–sulfur protein biogenesis

Banci

Bertini

Calderone

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Biogenesis of iron-sulfur cluster proteins is a highly regulated process that requires complex protein machineries. In the cytosolic iron-sulfur protein assembly machinery, two human key proteins-NADPH-dependent diflavin oxidoreductase 1 (Ndor1) and anamorsinform a stable complex in vivo that was proposed to provide electrons for assembling cytosolic iron-sulfur cluster proteins. The Ndor1-anamorsin interaction was also suggested to be implicated in the regulation of cell survival/death mechanisms. In the present work we unravel the molecular basis of recognition between Ndor1 and anamorsin and of the electron transfer process. This is based on the structural characterization of the two partner proteins, the investigation of the electron transfer process, and the identification of those protein regions involved in complex formation and those involved in electron transfer. We found that an unstructured region of anamorsin is essential for the formation of a specific and stable protein complex with Ndor1, whereas the C-terminal region of anamorsin, containing the [2Fe-2S] redox center, transiently interacts through complementary charged residues with the FMN-binding site region of Ndor1 to perform electron transfer. Our results propose a molecular model of the electron transfer process that is crucial for understanding the functional role of this interaction in human cells.Fe/S protein maturation | CIAPIN1 domain | diflavin reductase I ron-sulfur clusters (ISCs) are ancient inorganic cofactors that are crucial for many protein functions in eukaryotic and bacterial cells (1-3). The clusters are composed of inorganic sulfide and ferric/ ferrous iron atoms, the latter being preferentially coordinated by cysteinyl residues (4-6). Because inorganic sulfide and ferrous/ferric iron atoms are toxic in vivo, biosynthesis of ISC proteins within cells is a highly regulated process that requires complex protein machineries for the mobilization of Fe and S atoms from appropriate sources, for their assembly into ISC forms and their final delivery to the recipient proteins (7-9). Three distinct protein machineries are operative and essential in the (nonplant) eukaryotic cells for the biogenesis of ISC proteins: (i) the ISC assembly machinery in the mitochondrial matrix, (ii) the ISC export machinery located in the mitochondrial intermembrane space, and (iii) the cytosolic iron-sulfur protein assembly (CIA) machinery.The CIA machinery comprises several proteins (10, 11), among which one named Dre2 has been recently identified in yeast (12). The C-terminal domain of Dre2 (residues 228-348) is able to bind two ISCs, a [2Fe-2S] and a [4Fe-4S] (12, 13). The [2Fe-2S] cluster of Dre2 receives electrons from a cytosolic diflavin reductase, termed Tah18 (13), which contains a FAD and a FMN prosthetic group, respectively, bound in two distinct structural domains, to accept electrons from NADPH (14). Dre2 and Tah18 are protein partners forming a stable complex in vivo (13, 15). The C-terminal region of Dre2 (residues 173-348) is fundamenta...

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Section: Discussionmentioning

confidence: 79%

mentioning

confidence: 96%

mentioning

confidence: 99%

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Molecular view of an electron transfer process essential for iron–sulfur protein biogenesis

Banci

Bertini

Calderone

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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“…S288C) (29). These strains are also hypersensitive to the Y ⅐ quenching reagent hydroxyurea (28). We have been able to take advantage of this viability and our ability to permeabilize WT and ⌬rnr4 cells to take up proteins (e.g.…”

Section: Ribonucleotide Reductase (Rnr)mentioning

confidence: 97%

“…␤Ј is structurally homologous to ␤ but lacks three iron ligands, and as a consequence, contains no metallo-cofactor (18,22,(25)(26)(27). Although ␤Ј is catalytically inactive, it is required for converting ␤ into a conformation that is competent for iron loading and Y ⅐ formation in vitro and in vivo (23,24,28). Importantly, recombinant apo-␤ 2 and ␤Ј 2 rapidly form apo-␤␤Ј in vitro (24), and although cluster assembly from Fe II , O 2 , and reductant is inefficient (0.25 Y ⅐ /␤␤Ј), these properties allow us to study reconstitution in vitro with mutant small subunits.…”

Section: Ribonucleotide Reductase (Rnr)mentioning

confidence: 99%

Investigation of in Vivo Roles of the C-terminal Tails of the Small Subunit (ββ′) of Saccharomyces cerevisiae Ribonucleotide Reductase

Zhang

Stubbe

et al. 2013

Journal of Biological Chemistry

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Background: S. cerevisiae ribonucleotide reductase (RNR) comprises ␣ and ␤␤Ј subunits in an (␣ 2 ) m (␤␤Ј) n active holoenzyme. Results: C-terminal tail mutants of ␤␤Ј prevent radical transfer across the ␣-␤ interface and consequently deoxynucleotide production in vivo. Conclusion:The ␤␤Ј C-terminal tails interact only with the proximal ␣ within each ␣/␤(␣/␤Ј) pair. Significance: The predisposition of the ␤␤Ј C-terminal tails in active RNR in vivo is established.

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FeSCluster Assembly: Role of MonothiolGrxsandNfuProteins

Przybyla-Toscano

Roret

Couturier

et al. 2017

Encyclopedia of Inorganic and Bioinorganic Chemistry

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The cellular maturation of iron–sulfur (Fe–S) proteins necessitates dedicated assembly machineries. In eukaryotes, the ISC (iron–sulfur cluster) and SUF (sulfur mobilization) machineries are autonomous biogenesis systems for mitochondrial and chloroplastic proteins respectively, whereas the cytosolic CIA (cytosolic iron–sulfur cluster assembly) machinery operates for cytosolic and nuclear proteins and depends on the ISC machinery for obtaining the required labile sulfur atoms. Owing to their capacity to incorporate Fe–S clusters alone or in complex with BOLA proteins, class II glutaredoxins (GRXs) play some key roles in this maturation process, although the precise roles have yet to be clearly delineated. Organellar GRXs likely serve as Fe–S transfer proteins, accepting preformed Fe–S clusters from scaffold proteins used for their de novo synthesis before delivering them to other ISC or SUF targeting factors such as ISCA proteins or to final client proteins. The cytosolic/nuclear localized GRXs seem to have at least three different crucial roles, likely originating from their key position connecting the ISC machinery to the cytosolic compartment. Indeed, the molecular analysis of yeast mutants suggests that it could be the primary recipient protein of the unknown sulfur compound exported from the mitochondria and which could serve to assemble its Fe–S cluster. The presence of the cluster seems mandatory for all identified functions, serving (i) for the maturation of Fe–S clusters in the CIA electron donor DRE2, (ii) for the regulation of iron homeostasis in conjunction with BOLA and the Aft1/2 transcriptional regulators, and (iii) for the proper distribution of iron and correct functioning of all iron‐dependent proteins. These roles might not all be evolutionary conserved, notably in plants. In addition to providing an overview of the known physiological roles of class II GRXs, we describe their biochemical and structural properties focusing in particular on their interaction with other maturation factors of the BOLA, DRE2, and ISCA families.

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Investigation of in Vivo Diferric Tyrosyl Radical Formation in Saccharomyces cerevisiae Rnr2 Protein

Cited by 49 publications

References 66 publications

Molecular view of an electron transfer process essential for iron–sulfur protein biogenesis

Molecular view of an electron transfer process essential for iron–sulfur protein biogenesis

Investigation of in Vivo Roles of the C-terminal Tails of the Small Subunit (ββ′) of Saccharomyces cerevisiae Ribonucleotide Reductase

FeSCluster Assembly: Role of MonothiolGrxsandNfuProteins

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