Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.1.3 (ATPase)
65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To clarify the key role of Rad50 in DNA double-strand break repair (DSBR), we biochemically and structurally characterized ATP-bound and ATP-free Rad50 catalytic domain (Rad50cd) from Pyrococcus furiosus. Rad50cd displays ATPase activity plus ATP-controlled dimerization and DNA binding activities. Rad50cd crystal structures identify probable protein and DNA interfaces and reveal an ABC-ATPase fold, linking Rad50 molecular mechanisms to ABC transporters, including P glycoprotein and cystic fibrosis transmembrane conductance regulator. Binding of ATP gamma-phosphates to conserved signature motifs in two opposing Rad50cd molecules promotes dimerization that likely couples ATP hydrolysis to dimer dissociation and DNA release. These results, validated by mutations, suggest unified molecular mechanisms for ATP-driven cooperativity and allosteric control of ABC-ATPases in DSBR, membrane transport, and chromosome condensation by SMC proteins.
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PMID:Structural biology of Rad50 ATPase: ATP-driven conformational control in DNA double-strand break repair and the ABC-ATPase superfamily. 1089 49

SMC (structural maintenance of chromosomes) proteins are large coiled-coil proteins involved in chromosome condensation, sister chromatid cohesion, and DNA double-strand break processing. They share a conserved five-domain architecture with three globular domains separated by two long coiled-coil segments. The coiled-coil segments are antiparallel, bringing the N and C-terminal globular domains together. We have expressed a fusion protein of the N and C-terminal globular domains of Thermotoga maritima SMC in Escherichia coli by replacing the approximately 900 residue coiled-coil and hinge segment with a short peptide linker. The SMC head domain (SMChd) binds and condenses DNA in an ATP-dependent manner. Using selenomethionine-substituted protein and multiple anomalous dispersion phasing, we have solved the crystal structure of the SMChd to 3.1 A resolution. In the monoclinic crystal form, six SMChd molecules form two turns of a helix. The fold of SMChd is closely related to the ATP-binding cassette (ABC) ATPase family of proteins and Rad50, a member of the SMC family involved in DNA double-strand break repair. In SMChd, the ABC ATPase fold is formed by the N and C-terminal domains with the 900 residue coiled-coil and hinge segment inserted in the middle of the fold. The crystal structure of an SMChd confirms that the coiled-coil segments in SMC proteins are anti-parallel and shows how the N and C-terminal domains come together to form an ABC ATPase. Comparison to the structure of the MukB N-terminal domain demonstrates the close relationship between MukB and SMC proteins, and indicates a helix to strand conversion when N and C-terminal parts come together.
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PMID:Crystal structure of the SMC head domain: an ABC ATPase with 900 residues antiparallel coiled-coil inserted. 1117 91

To clarify functions of the Mre11/Rad50 (MR) complex in DNA double-strand break repair, we report Pyrococcus furiosus Mre11 crystal structures, revealing a protein phosphatase-like, dimanganese binding domain capped by a unique domain controlling active site access. These structures unify Mre11's multiple nuclease activities in a single endo/exonuclease mechanism and reveal eukaryotic macromolecular interaction sites by mapping human and yeast Mre11 mutations. Furthermore, the structure of the P. furiosus Rad50 ABC-ATPase with its adjacent coiled-coil defines a compact Mre11/Rad50-ATPase complex and suggests that Rad50-ATP-driven conformational switching directly controls the Mre11 exonuclease. Electron microscopy, small angle X-ray scattering, and ultracentrifugation data of human and P. furiosus MR reveal a dual functional complex consisting of a (Mre11)2/(Rad50)2 heterotetrameric DNA processing head and a double coiled-coil linker.
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PMID:Structural biochemistry and interaction architecture of the DNA double-strand break repair Mre11 nuclease and Rad50-ATPase. 1137 44

The RAD50 gene of Saccharomyces cerevisiae is one of several genes required for recombinational repair of double-strand DNA breaks during vegetative growth and for initiation of meiotic recombination. Rad50 forms a complex with two other proteins, Mre11 and Xrs2, and this complex is involved in double-strand break formation and processing. Rad50 has limited sequence homology to the structural maintenance of chromosomes (SMC) family of proteins and shares the same domain structure as SMCs: N- and C-terminal globular domains separated by two long coiled-coils. However, a notable difference is the much smaller non-coil hinge region between the two coiled-coils. We report here a structural analysis of full-length S. cerevisiae Rad50, alone and in a complex with yeast Mre11 by electron microscopy. Our results confirm that yeast Rad50 does have the same antiparallel coiled-coil structure as SMC proteins, but with no detectable globular hinge domain. However, the molecule is still able to bend sharply in the middle to bring the two catalytic domains together, indicating that the small hinge domain is flexible. We also demonstrate that Mre11 binds as a dimer between the catalytic domains of Rad50, bringing the nuclease activities of Mre11 in close proximity to the ATPase and DNA binding activities of Rad50.
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PMID:Structure of the Rad50 x Mre11 DNA repair complex from Saccharomyces cerevisiae by electron microscopy. 1147 Aug

Mre11-Rad50 (MR) proteins are encoded by bacteriophage, eubacterial, archeabacterial and eukaryotic genomes, and form a complex with a remarkable protein architecture. This complex is capable of tethering the ends of DNA molecules, possesses a variety of DNA nuclease, helicase, ATPase and annealing activities, and performs a wide range of functions within cells. It is required for meiotic recombination, double-strand break repair, processing of mis-folded DNA structures and maintaining telomere length. This article reviews current knowledge of the structure and enzymatic activities of the MR complex and attempts to integrate biochemical information with the roles of the protein in a cell.
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PMID:Tethering on the brink: the evolutionarily conserved Mre11-Rad50 complex. 1215 Dec 26

The Mre11 complex (Mre11 Rad50 Nbs1) is central to chromosomal maintenance and functions in homologous recombination, telomere maintenance and sister chromatid association. These functions all imply that the linked binding of two DNA substrates occurs, although the molecular basis for this process remains unknown. Here we present a 2.2 A crystal structure of the Rad50 coiled-coil region that reveals an unexpected dimer interface at the apex of the coiled coils in which pairs of conserved Cys-X-X-Cys motifs form interlocking hooks that bind one Zn(2+) ion. Biochemical, X-ray and electron microscopy data indicate that these hooks can join oppositely protruding Rad50 coiled-coil domains to form a flexible bridge of up to 1,200 A. This suggests a function for the long insertion in the Rad50 ABC-ATPase domain. The Rad50 hook is functional, because mutations in this motif confer radiation sensitivity in yeast and disrupt binding at the distant Mre11 nuclease interface. These data support an architectural role for the Rad50 coiled coils in forming metal-mediated bridging complexes between two DNA-binding heads. The resulting assemblies have appropriate lengths and conformational properties to link sister chromatids in homologous recombination and DNA ends in non-homologous end-joining.
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PMID:The Rad50 zinc-hook is a structure joining Mre11 complexes in DNA recombination and repair. 1215 85

The present report deals with the functional relationships among protein complexes which, when mutated, are responsible for four human syndromes displaying cancer proneness, and whose cells are deficient in DNA double-strand break (DSB) repair. In some of them, the cells are also unable to activate the proper checkpoint, while in the others an unduly override of the checkpoint-induced arrest occurs. As a consequence, all these patients display genome instability. In ataxia-telangiectasia, the mutated protein (ATM) is a kinase, which acts as a transducer of DNA damage signalling. The defective protein in the ataxia-telangiectasia-like disorder is a DNase (the Mre11 nuclease) that in vivo produces single-strand tails at both sides of DSBs. Mre11 is always present with the Rad50 ATPase in a protein machine: the nuclease complex. In mammals, this complex also contains nibrin, the protein mutated in the Nijmegen syndrome. Nibrin confers new abilities to the nuclease complex, and can also bind to BRCA1 (one of the two proteins mutated in familial breast cancer). BRCA1 has a central motif that binds with high affinity to cruciform DNA, a structure present in places where the DNA loops are anchored to the chromosomal axis or scaffold. The BRCA1 x cruciform DNA complex should be released to allow the nuclease complex to work in DNA recombinational repair of DSBs. BRCA1 also acts as a scaffold for the assembly of ATPases such as Rad51, responsible for the somatic homologous recombination. Loss of the BRCA1 gene prevents cell survival after exposure to cross-linkers. The BRCA1-RING domain is an E3-ubiquitin ligase. It can mono-ubiquitinate the FANCD2 protein, mutated in one of the Fanconi anemia complementation groups, to regulate it. Finally, during DNA replication, the nuclease complex and its activating ATM kinase are integrated in the BRCA1-associated surveillance complex (BASC) that contains, among others, enzymes required for mismatch excision repair. In short, the proteins missing in these syndromes have in common their BRCA1-mediated assembly into multimeric machines responsible for the surveillance of DNA replication, DSB recombinational repair, and the removal of DNA cross-links.
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PMID:Human syndromes with genomic instability and multiprotein machines that repair DNA double-strand breaks. 1250 2

We identify and characterize MlaA, a novel protein, which is found in a conserved operon with Mre11 and Rad50 in archaeal genomes. MlaA is fused with Mre11 in Methanobacter thermoautotrophicus, suggesting the MlaA is functionally linked to the Mre11 complex. MlaA preferentially and cooperatively binds double-stranded and secondary structure containing DNA and has double-stranded but not single-stranded DNA-stimulated ATPase activity. Electron microscopy reveals that MlaA forms a 360-kDa hexameric ring structure with a central hole. Our data suggest that the archaeal Mre11 complex is associated with a novel hexameric ATPase that could be required for the processing of DNA double-stranded breaks and recombination intermediates.
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PMID:MlaA, a hexameric ATPase linked to the Mre11 complex in archaeal genomes. 1471 Jan 87

Structural maintenance of chromosomes (SMC) proteins have diverse cellular functions including chromosome segregation, condensation and DNA repair. They are grouped based on a conserved set of distinct structural motifs. All SMC proteins are predicted to have a bipartite ATPase domain that is separated by a long region predicted to form a coiled coil. Recent structural data on a variety of SMC proteins shows them to be arranged as long intramolecular coiled coils with a globular ATPase at one end. SMC proteins function in pairs as heterodimers or as homodimers often in complexes with other proteins. We expect the arrangement of the SMC protein domains in complex assemblies to have important implications for their diverse functions. We used scanning force microscopy imaging to determine the architecture of human, Saccharomyces cerevisiae, and Pyrococcus furiosus Rad50/Mre11, Escherichia coli SbcCD, and S.cerevisiae SMC1/SMC3 cohesin SMC complexes. Two distinct architectural arrangements are described, based on the way their components were connected. The eukaryotic complexes were similar to each other and differed from their prokaryotic and archaeal homologs. These similarities and differences are discussed with respect to their diverse mechanistic roles in chromosome metabolism.
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PMID:Differential arrangements of conserved building blocks among homologs of the Rad50/Mre11 DNA repair protein complex. 1516 61

Recently, it has been shown that a predicted P-loop ATPase (the HerA or MlaA protein), which is highly conserved in archaea and also present in many bacteria but absent in eukaryotes, has a bidirectional helicase activity and forms hexameric rings similar to those described for the TrwB ATPase. In this study, the FtsK-HerA superfamily of P-loop ATPases, in which the HerA clade comprises one of the major branches, is analyzed in detail. We show that, in addition to the FtsK and HerA clades, this superfamily includes several families of characterized or predicted ATPases which are predominantly involved in extrusion of DNA and peptides through membrane pores. The DNA-packaging ATPases of various bacteriophages and eukaryotic double-stranded DNA viruses also belong to the FtsK-HerA superfamily. The FtsK protein is the essential bacterial ATPase that is responsible for the correct segregation of daughter chromosomes during cell division. The structural and evolutionary relationship between HerA and FtsK and the nearly perfect complementarity of their phyletic distributions suggest that HerA similarly mediates DNA pumping into the progeny cells during archaeal cell division. It appears likely that the HerA and FtsK families diverged concomitantly with the archaeal-bacterial division and that the last universal common ancestor of modern life forms had an ancestral DNA-pumping ATPase that gave rise to these families. Furthermore, the relationship of these cellular proteins with the packaging ATPases of diverse DNA viruses suggests that a common DNA pumping mechanism might be operational in both cellular and viral genome segregation. The herA gene forms a highly conserved operon with the gene for the NurA nuclease and, in many archaea, also with the orthologs of eukaryotic double-strand break repair proteins MRE11 and Rad50. HerA is predicted to function in a complex with these proteins in DNA pumping and repair of double-stranded breaks introduced during this process and, possibly, also during DNA replication. Extensive comparative analysis of the 'genomic context' combined with in-depth sequence analysis led to the prediction of numerous previously unnoticed nucleases of the NurA superfamily, including a specific version that is likely to be the endonuclease component of a novel restriction-modification system. This analysis also led to the identification of previously uncharacterized nucleases, such as a novel predicted nuclease of the Sir2-type Rossmann fold, and phosphatases of the HAD superfamily that are likely to function as partners of the FtsK-HerA superfamily ATPases.
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PMID:Comparative genomics of the FtsK-HerA superfamily of pumping ATPases: implications for the origins of chromosome segregation, cell division and viral capsid packaging. 1546 93


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