scholarly journals Structural Basis of RICs Iron Donation for Iron-Sulfur Cluster Biogenesis

2021 ◽  
Vol 12 ◽  
Author(s):  
Liliana S. O. Silva ◽  
Pedro M. Matias ◽  
Célia V. Romão ◽  
Lígia M. Saraiva
Keyword(s):  
Protein Interactions ◽  
Structural Basis ◽  
Single Mutation ◽  
Protein Protein Interactions ◽  
Cysteine Desulfurase ◽  
Iron Sulfur Cluster ◽  
Sulfur Cluster ◽  
E Coli ◽  
Iron Sulfur ◽  
Crucial Component

Escherichia coli YtfE is a di-iron protein of the widespread Repair of Iron Centers proteins (RIC) family that has the capacity to donate iron, which is a crucial component of the biogenesis of the ubiquitous family of iron-sulfur proteins. In this work we identify in E. coli a previously unrecognized link between the YtfE protein and the major bacterial system for iron-sulfur cluster (ISC) assembly. We show that YtfE establishes protein-protein interactions with the scaffold IscU, where the transient cluster is formed, and the cysteine desulfurase IscS. Moreover, we found that promotion by YtfE of the formation of an Fe-S cluster in IscU requires two glutamates, E125 and E159 in YtfE. Both glutamates form part of the entrance of a protein channel in YtfE that links the di-iron center to the surface. In particular, E125 is crucial for the exit of iron, as a single mutation to leucine closes the channel rendering YtfE inactive for the build-up of Fe-S clusters. Hence, we provide evidence for the key role of RICs as bacterial iron donor proteins involved in the biogenesis of Fe-S clusters.

scholarly journals Structural basis of RICs iron donation for iron-sulfur cluster biogenesis

2021 ◽  
Author(s):  
Liliana S. O. Silva ◽  
Pedro M. Matias ◽  
Célia V. Romão ◽  
Lígia M. Saraiva
Keyword(s):  
Escherichia Coli ◽  
Protein Interactions ◽  
Structural Characteristics ◽  
Structural Basis ◽  
Single Mutation ◽  
Protein Protein Interactions ◽  
Iron Sulfur Cluster ◽  
Sulfur Cluster ◽  
Iron Sulfur ◽  
Iron Sulfur Proteins

AbstractEscherichia coli YtfE is a di-iron protein, of the widespread RIC family, with capacity to donate iron, which is a crucial component of the biogenesis of the ubiquitous family of iron-sulfur proteins. Herein we identify in E. coli a previously unrecognized link between the YtfE protein and the major bacterial system for iron-sulfur cluster (ISC) assembly. We show that YtfE establishes protein-protein interactions with the scaffold IscU, where the transient cluster is formed, and the cysteine desulfurase IscS. Moreover, we found that promotion by YtfE of the formation of an Fe-S cluster in IscU requires two glutamates, E125 and E159 in YtfE. Both glutamates form part of the entrance of a protein channel in YtfE that links the di-iron centre to the surface. In particular, E125 is crucial for the exit of iron, as a single mutation to leucine closes the channel rendering YtfE inactive for the build-up of Fe-S clusters. Hence, we provide evidence for the key role of RICs as bacterial iron donor proteins involved in the biogenesis of Fe-S clusters.ImportanceThe ubiquitous iron-sulfur proteins require specialized cellular machineries to promote the assembly of the cofactor. These systems include proteins that provide sulfur and iron, and scaffold proteins where the cluster is formed. Although largely studied the nature of the iron donor remains to be fully clarified. In this work, we show that Escherichia coli YtfE, which belongs to the RIC protein family, establishes protein-protein interactions with two of the major proteins of the ISC system, and we reveal the structural characteristics necessary for the exit of iron ions from YtfE. Altogether our results prove that RICs can be considered a family of iron donor proteins involved in the biogenesis of iron-sulfur containing proteins.

scholarly journals New Techniques for Ancient Proteins: Direct Coupling Analysis Applied on Proteins involved in Iron Sulfur Cluster Biogenesis

2017 ◽  
Author(s):  
Marco Fantini ◽  
Duccio Malinverni ◽  
Paolo De Los Rios ◽  
Annalisa Pastore
Keyword(s):  
Protein Interactions ◽  
Strong Support ◽  
Scaffold Protein ◽  
Direct Coupling ◽  
Protein Protein Interactions ◽  
Coupling Analysis ◽  
Iron Sulfur Cluster ◽  
Sulfur Cluster ◽  
Iron Sulfur ◽  
Direct Coupling Analysis

ABSTRACTDirect coupling analysis (DCA) is a powerful tool based on protein evolution and introduced to predict protein fold and protein-protein interactions which has been applied also to the prediction of entire interactomes. We have used DCA to analyse three proteins of the iron-sulfur biogenesis machine, an essential metabolic pathway conserved in all organisms. We show that, although based on a relatively small number of sequences due to its distribution in genomes, we can correctly recapitulate all the features of the fold of the CyaY/frataxin family, a protein involved in the human disease Friedreich’s ataxia. This result gave us confidence in the use of this tool. Application of DCA to the iron-sulfur cluster scaffold protein IscU, which has been suggested to function both as an ordered and a disordered form, allows us to clearly distinguish evolutionary traces of the structured species, suggesting that, if present in the cell, the disordered form has not left any evolutionary imprinting. We observe instead, for the first time, direct indications of how the protein can dimerize head-to-head and bind 4Fe4S clusters. Analysis of the alternative scaffold protein IscA provides strong support to a coordination of the cluster mediated by a dimeric rather than a tetrameric form as previously suggested. Our analysis also suggests the presence in solution of a mixture of monomeric and dimeric species and guide us to the prevalent one. Finally, we used DCA to analyse protein-protein interactions between some of these proteins and discuss the potentialities and the limitations of the method.

scholarly journals Functional Studies of [FeFe] Hydrogenase Maturation in an Escherichia coli Biosynthetic System

2006 ◽  
Vol 188 (6) ◽  
pp. 2163-2172 ◽  
Author(s):  
Paul W. King ◽  
Matthew C. Posewitz ◽  
Maria L. Ghirardi ◽  
Michael Seibert
Keyword(s):  
Escherichia Coli ◽  
Catalytic Domain ◽  
Expression System ◽  
Iron Sulfur Cluster ◽  
Sulfur Cluster ◽  
E Coli ◽  
Functional Studies ◽  
Hydrogenase Maturation ◽  
Iron Sulfur ◽  
And Function

ABSTRACT Maturation of [FeFe] hydrogenases requires the biosynthesis and insertion of the catalytic iron-sulfur cluster, the H cluster. Two radical S-adenosylmethionine (SAM) proteins proposed to function in H cluster biosynthesis, HydEF and HydG, were recently identified in the hydEF-1 mutant of the green alga Chlamydomonas reinhardtii (M. C. Posewitz, P. W. King, S. L. Smolinski, L. Zhang, M. Seibert, and M. L. Ghirardi, J. Biol. Chem. 279:25711-25720, 2004). Previous efforts to study [FeFe] hydrogenase maturation in Escherichia coli by coexpression of C. reinhardtii HydEF and HydG and the HydA1 [FeFe] hydrogenase were hindered by instability of the hydEF and hydG expression clones. A more stable [FeFe] hydrogenase expression system has been achieved in E. coli by cloning and coexpression of hydE, hydF, and hydG from the bacterium Clostridium acetobutylicum. Coexpression of the C. acetobutylicum maturation proteins with various algal and bacterial [FeFe] hydrogenases in E. coli resulted in purified enzymes with specific activities that were similar to those of the enzymes purified from native sources. In the case of structurally complex [FeFe] hydrogenases, maturation of the catalytic sites could occur in the absence of an accessory iron-sulfur cluster domain. Initial investigations of the structure and function of the maturation proteins HydE, HydF, and HydG showed that the highly conserved radical-SAM domains of both HydE and HydG and the GTPase domain of HydF were essential for achieving biosynthesis of active [FeFe] hydrogenases. Together, these results demonstrate that the catalytic domain and a functionally complete set of Hyd maturation proteins are fundamental to achieving biosynthesis of catalytic [FeFe] hydrogenases.

scholarly journals Zinc Toxicity and Iron-Sulfur Cluster Biogenesis inEscherichia coli

2019 ◽  
Vol 85 (9) ◽  
Author(s):  
Jianghui Li ◽  
Xiaojun Ren ◽  
Bingqian Fan ◽  
Zhaoyang Huang ◽  
Wu Wang ◽  
...  
Keyword(s):  
Binding Activity ◽  
Zinc Toxicity ◽  
Cluster Assembly ◽  
Intracellular Zinc ◽  
Iron Sulfur Cluster ◽  
Content Type ◽  
Sulfur Cluster ◽  
E Coli ◽  
Iron Sulfur ◽  
Assembly Proteins

ABSTRACTWhile zinc is an essential trace metal in biology, excess zinc is toxic to organisms. Previous studies have shown that zinc toxicity is associated with disruption of the [4Fe-4S] clusters in various dehydratases inEscherichia coli. Here, we report that the intracellular zinc overload inE. colicells inhibits iron-sulfur cluster biogenesis without affecting the preassembled iron-sulfur clusters in proteins. Among the housekeeping iron-sulfur cluster assembly proteins encoded by the gene clusteriscSUA-hscBA-fdx-iscXinE. colicells, the scaffold IscU, the iron chaperone IscA, and ferredoxin have strong zinc binding activity in cells, suggesting that intracellular zinc overload inhibits iron-sulfur cluster biogenesis by binding to the iron-sulfur cluster assembly proteins. Mutations of the conserved cysteine residues to serine in IscA, IscU, or ferredoxin completely abolish the zinc binding activity of the proteins, indicating that zinc can compete with iron or iron-sulfur cluster binding in IscA, IscU, and ferredoxin and block iron-sulfur cluster biogenesis. Furthermore, intracellular zinc overload appears to emulate the slow-growth phenotype of theE. colimutant cells with deletion of the iron-sulfur cluster assembly proteins IscU, IscA, and ferredoxin. Our results suggest that intracellular zinc overload inhibits iron-sulfur cluster biogenesis by targeting the iron-sulfur cluster assembly proteins IscU, IscA, and ferredoxin inE. colicells.IMPORTANCEZinc toxicity has been implicated in causing various human diseases. High concentrations of zinc can also inhibit bacterial cell growth. However, the underlying mechanism has not been fully understood. Here, we report that zinc overload inEscherichia colicells inhibits iron-sulfur cluster biogenesis by targeting specific iron-sulfur cluster assembly proteins. Because iron-sulfur proteins are involved in diverse physiological processes, the zinc-mediated inhibition of iron-sulfur cluster biogenesis could be largely responsible for the zinc-mediated cytotoxicity. Our finding provides new insights on how intracellular zinc overload may inhibit cellular functions in bacteria.

scholarly journals The Role of CyaY in Iron Sulfur Cluster Assembly on the E. coli IscU Scaffold Protein

PLoS ONE ◽  
2011 ◽  
Vol 6 (7) ◽  
pp. e21992 ◽  
Author(s):  
Clara Iannuzzi ◽  
Salvatore Adinolfi ◽  
Barry D. Howes ◽  
Ricardo Garcia-Serres ◽  
Martin Clémancey ◽  
...  
Keyword(s):  
Scaffold Protein ◽  
Cluster Assembly ◽  
Iron Sulfur Cluster ◽  
Sulfur Cluster ◽  
E Coli ◽  
Iron Sulfur
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