Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0016719 (Friedreich's ataxia)
 2,098 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Friedreich's ataxia (FRDA) is a progressive neurodegenerative disease associated with the loss of function of the protein frataxin (FXN) that results from low FXN levels due to a GAA triplet repeat expansion or, occasionally, from missense mutations in the FXN gene. Here biochemical and structural properties of FXN variants, including three FRDA missense mutations (N146K, Q148R, and R165C) and three related mutants (N146A, Q148G, and Q153A), were determined in an effort to understand the structural basis for the loss of function. In vitro assays revealed that although the three FRDA missense mutations exhibited similar losses of cysteine desulfurase and Fe-S cluster assembly activities, the causes for these activation defects were distinct. The R165C variant exhibited a k(cat)/K(M) higher than that of native FXN but weak binding to the NFS1, ISD11, and ISCU2 (SDU) complex, whereas the Q148R variant exhibited the lowest k(cat)/K(M) of the six tested FXN variants and only a modest binding deficiency. The order of the FXN binding affinities for the SDU Fe-S assembly complex was as follows: FXN > Q148R > N146A > Q148G > N146K > Q153A > R165C. Four different classes of FXN variants were identified on the basis of their biochemical properties. Together, these structure-function studies reveal determinants for the binding and allosteric activation of the Fe-S assembly complex and provide insight into how FRDA missense mutations are functionally compromised.
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PMID:Structure-function analysis of Friedreich's ataxia mutants reveals determinants of frataxin binding and activation of the Fe-S assembly complex. 2177 84

Human frataxin (FXN) has been intensively studied since the discovery that the FXN gene is associated with the neurodegenerative disease Friedreich's ataxia. Human FXN is a component of the NFS1-ISD11-ISCU2-FXN (SDUF) core Fe-S assembly complex and activates the cysteine desulfurase and Fe-S cluster biosynthesis reactions. In contrast, the Escherichia coli FXN homologue CyaY inhibits Fe-S cluster biosynthesis. To resolve this discrepancy, enzyme kinetic experiments were performed for the human and E. coli systems in which analogous cysteine desulfurase, Fe-S assembly scaffold, and frataxin components were interchanged. Surprisingly, our results reveal that activation or inhibition by the frataxin homologue is determined by which cysteine desulfurase is present and not by the identity of the frataxin homologue. These data are consistent with a model in which the frataxin-less Fe-S assembly complex exists as a mixture of functional and nonfunctional states, which are stabilized by binding of frataxin homologues. Intriguingly, this appears to be an unusual example in which modifications to an enzyme during evolution inverts or reverses the mode of control imparted by a regulatory molecule.
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PMID:Effector role reversal during evolution: the case of frataxin in Fe-S cluster biosynthesis. 2235 84

Iron-sulfur (Fe-S) cluster-containing proteins are essential components of cells. In eukaryotes, Fe-S clusters are synthesized by the mitochondrial iron-sulfur cluster (ISC) machinery and the cytosolic iron-sulfur assembly (CIA) system. In the mammalian ISC machinery, preassembly of the Fe-S cluster on the scaffold protein (ISCU) involves a cysteine desulfurase complex (NFS1/ISD11) and frataxin (FXN), the protein deficient in Friedreich's ataxia. Here, by comparing the biochemical and spectroscopic properties of quaternary (ISCU/NFS1/ISD11/FXN) and ternary (ISCU/NFS1/ISD11) complexes, we show that FXN stabilizes the quaternary complex and controls iron entry to the complex through activation of cysteine desulfurization. Furthermore, we show for the first time that in the presence of iron and L-cysteine, an [Fe(4)S(4)] cluster is formed within the quaternary complex that can be transferred to mammalian aconitase (mACO2) to generate an active enzyme. In the absence of FXN, although the ternary complex can assemble an Fe-S cluster, the cluster is inefficiently transferred to ACO2. Taken together, these data help to unravel further the Fe-S cluster assembly process and the molecular basis of Friedreich's ataxia.
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PMID:Mammalian frataxin controls sulfur production and iron entry during de novo Fe4S4 cluster assembly. 2326 91

Iron-sulfur (Fe-S) clusters are a class of highly conserved and ubiquitous prosthetic groups with unique chemical properties that allow the proteins that contain them, Fe-S proteins, to assist in various key biochemical pathways. Mutations in Fe-S proteins often disrupt Fe-S cluster assembly leading to a spectrum of severe disorders such as Friedreich's ataxia or iron-sulfur cluster assembly enzyme (ISCU) myopathy. Herein, we describe infantile mitochondrial complex II/III deficiency, a novel autosomal recessive mitochondrial disease characterized by lactic acidemia, hypotonia, respiratory chain complex II and III deficiency, multisystem organ failure and abnormal mitochondria. Through autozygosity mapping, exome sequencing, in silico analyses, population studies and functional tests, we identified c.215G>A, p.Arg72Gln in NFS1 as the likely causative mutation. We describe the first disease in man likely caused by deficiency in NFS1, a cysteine desulfurase that is implicated in respiratory chain function and iron maintenance by initiating Fe-S cluster biosynthesis. Our results further demonstrate the importance of sufficient NFS1 expression in human physiology.
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PMID:Exome sequencing identifies NFS1 deficiency in a novel Fe-S cluster disease, infantile mitochondrial complex II/III deficiency. 2449 31

Iron-sulfur clusters are ubiquitous protein cofactors with critical cellular functions. The mitochondrial Fe-S assembly complex, which consists of the cysteine desulfurase NFS1 and its accessory protein (ISD11), the Fe-S assembly protein (ISCU2), and frataxin (FXN), converts substrates l-cysteine, ferrous iron, and electrons into Fe-S clusters. The physiological function of FXN has received a tremendous amount of attention since the discovery that its loss is directly linked to the neurodegenerative disease Friedreich's ataxia. Previous in vitro results revealed a role for human FXN in activating the cysteine desulfurase and Fe-S cluster biosynthesis activities of the Fe-S assembly complex. Here we present radiolabeling experiments that indicate FXN accelerates the accumulation of sulfur on ISCU2 and that the resulting persulfide species is viable in the subsequent synthesis of Fe-S clusters. Additional mutagenesis, enzyme kinetic, UV-visible, and circular dichroism spectroscopic studies suggest conserved ISCU2 residue C104 is critical for FXN activation, whereas C35, C61, and C104 are all essential for Fe-S cluster formation on the assembly complex. These results cannot be fully explained by the hypothesis that FXN functions as an iron donor for Fe-S cluster biosynthesis, and further support an allosteric regulator role for FXN. Together, these results lead to an activation model in which FXN accelerates persulfide formation on NFS1 and favors a helix-to-coil interconversion on ISCU2 that facilitates the transfer of sulfur from NFS1 to ISCU2 as an initial step in Fe-S cluster biosynthesis.
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PMID:Human frataxin activates Fe-S cluster biosynthesis by facilitating sulfur transfer chemistry. 2497 90

Iron-sulfur (Fe-S) clusters are essential cofactors, and mitochondria contain several Fe-S proteins, including the [4Fe-4S] protein aconitase and the [2Fe-2S] protein ferredoxin. Fe-S cluster assembly of these proteins occurs within mitochondria. Although considerable data exist for yeast mitochondria, this biosynthetic process has never been directly demonstrated in mammalian mitochondria. Using [(35)S]cysteine as the source of sulfur, here we show that mitochondria isolated from Cath.A-derived cells, a murine neuronal cell line, can synthesize and insert new Fe-(35)S clusters into aconitase and ferredoxins. The process requires GTP, NADH, ATP, and iron, and hydrolysis of both GTP and ATP is necessary. Importantly, we have identified the (35)S-labeled persulfide on the NFS1 cysteine desulfurase as a genuine intermediate en route to Fe-S cluster synthesis. In physiological settings, the persulfide sulfur is released from NFS1 and transferred to a scaffold protein, where it combines with iron to form an Fe-S cluster intermediate. We found that the release of persulfide sulfur from NFS1 requires iron, showing that the use of iron and sulfur for the synthesis of Fe-S cluster intermediates is a highly coordinated process. The release of persulfide sulfur also requires GTP and NADH, probably mediated by a GTPase and a reductase, respectively. ATP, a cofactor for a multifunctional Hsp70 chaperone, is not required at this step. The experimental system described here may help to define the biochemical basis of diseases that are associated with impaired Fe-S cluster biogenesis in mitochondria, such as Friedreich ataxia.
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PMID:Fe-S cluster biogenesis in isolated mammalian mitochondria: coordinated use of persulfide sulfur and iron and requirements for GTP, NADH, and ATP. 2539 79

Friedreich's ataxia is a severe neurodegenerative disease caused by the decreased expression of frataxin, a mitochondrial protein that stimulates iron-sulfur (Fe-S) cluster biogenesis. In mammals, the primary steps of Fe-S cluster assembly are performed by the NFS1-ISD11-ISCU complex via the formation of a persulfide intermediate on NFS1. Here we show that frataxin modulates the reactivity of NFS1 persulfide with thiols. We use maleimide-peptide compounds along with mass spectrometry to probe cysteine-persulfide in NFS1 and ISCU. Our data reveal that in the presence of ISCU, frataxin enhances the rate of two similar reactions on NFS1 persulfide: sulfur transfer to ISCU leading to the accumulation of a persulfide on the cysteine C104 of ISCU, and sulfur transfer to small thiols such as DTT, L-cysteine and GSH leading to persulfuration of these thiols and ultimately sulfide release. These data raise important questions on the physiological mechanism of Fe-S cluster assembly and point to a unique function of frataxin as an enhancer of sulfur transfer within the NFS1-ISD11-ISCU complex.
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PMID:Mammalian frataxin directly enhances sulfur transfer of NFS1 persulfide to both ISCU and free thiols. 2559 3

In eukaryotes, sulfur is mobilized for incorporation into multiple biosynthetic pathways by a cysteine desulfurase complex that consists of a catalytic subunit (NFS1), LYR protein (ISD11), and acyl carrier protein (ACP). This NFS1-ISD11-ACP (SDA) complex forms the core of the iron-sulfur (Fe-S) assembly complex and associates with assembly proteins ISCU2, frataxin (FXN), and ferredoxin to synthesize Fe-S clusters. Here we present crystallographic and electron microscopic structures of the SDA complex coupled to enzyme kinetic and cell-based studies to provide structure-function properties of a mitochondrial cysteine desulfurase. Unlike prokaryotic cysteine desulfurases, the SDA structure adopts an unexpected architecture in which a pair of ISD11 subunits form the dimeric core of the SDA complex, which clarifies the critical role of ISD11 in eukaryotic assemblies. The different quaternary structure results in an incompletely formed substrate channel and solvent-exposed pyridoxal 5'-phosphate cofactor and provides a rationale for the allosteric activator function of FXN in eukaryotic systems. The structure also reveals the 4'-phosphopantetheine-conjugated acyl-group of ACP occupies the hydrophobic core of ISD11, explaining the basis of ACP stabilization. The unexpected architecture for the SDA complex provides a framework for understanding interactions with acceptor proteins for sulfur-containing biosynthetic pathways, elucidating mechanistic details of eukaryotic Fe-S cluster biosynthesis, and clarifying how defects in Fe-S cluster assembly lead to diseases such as Friedreich's ataxia. Moreover, our results support a lock-and-key model in which LYR proteins associate with acyl-ACP as a mechanism for fatty acid biosynthesis to coordinate the expression, Fe-S cofactor maturation, and activity of the respiratory complexes.
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PMID:Structure of human Fe-S assembly subcomplex reveals unexpected cysteine desulfurase architecture and acyl-ACP-ISD11 interactions. 2863 2

Frataxin is the protein responsible for the genetically-inherited neurodegenerative disease Friedreich's ataxia caused by partial silencing of the protein and loss of function. Although the frataxin function is not yet entirely clear, it has been associated to the machine that builds iron-sulfur clusters, essential prosthetic groups involved in several processes and is strongly conserved in organisms from bacteria to humans. Two of its important molecular partners are the protein NFS1 (or IscS in bacteria), that is the desulfurase which converts cysteine to alanine and produces sulfur, and ISU (or IscU), the scaffold protein which transiently accepts the cluster. While bacterial frataxin has been extensively characterized, only few eukaryotic frataxins have been described. Here we report the 1H, 13C and 15N backbone and side-chain chemical shift assignments of frataxin from Chaetomium thermophilum, a thermophile increasingly used by virtue of its stability.
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PMID:Chemical shift assignment of a thermophile frataxin. 2909 Apr 18

Friedreich's ataxia is a disease caused by a decrease in the levels of expression or loss of functionality of the mitochondrial protein frataxin (FXN). The development of an active and stable recombinant variant of FXN is important for protein replacement therapy. Although valuable data about the mature form FXN81-210 has been collected, not enough information is available about the conformation of the frataxin precursor (FXN1-210). We investigated the conformation, stability and function of a recombinant precursor variant (His6-TAT-FXN1-210), which includes a TAT peptide in the N-terminal region to assist with transport across cell membranes. His6-TAT-FXN1-210 was expressed in Escherichia coli and conditions were found for purifying folded protein free of aggregation, oxidation or degradation, even after freezing and thawing. The protein was found to be stable and monomeric, with the N-terminal stretch (residues 1-89) mostly unstructured and the C-terminal domain properly folded. The experimental data suggest a complex picture for the folding process of full-length frataxin in vitro: the presence of the N-terminal region increased the tendency of FXN to aggregate at high temperatures but this could be avoided by the addition of low concentrations of GdmCl. The purified precursor was translocated through cell membranes. In addition, immune response against His6-TAT-FXN1-210 was measured, suggesting that the C-terminal fragment was not immunogenic at the assayed protein concentrations. Finally, the recognition of recombinant FXN by cellular proteins was studied to evaluate its functionality. In this regard, cysteine desulfurase NFS1/ISD11/ISCU was activated in vitro by His6-TAT-FXN1-210. Moreover, the results showed that His6-TAT-FXN1-210 can be ubiquitinated in vitro by the recently identified frataxin E3 ligase RNF126, in a similar way as the FXN1-210, suggesting that the His6-TAT extension does not interfere with the ubiquitination machinery.
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PMID:Biophysical characterisation of the recombinant human frataxin precursor. 2951 16


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