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)

Werner syndrome (WS) is a genetic premature aging disorder in which patients appear much older than their chronological age. The gene mutated in WS encodes a nuclear protein (WRN) which possesses 3'-5' exonuclease and ATPase-dependent 3'-5' helicase activities. The genomic instability associated with WS cells and the biochemical characteristics of WRN suggest that WRN plays a role in DNA metabolic pathways such as transcription, replication, recombination and repair. Recently we have identified poly(ADP-ribose) polymerase-1 (PARP-1) as a new WRN interacting protein. In this paper, we further mapped the interacting domains. We found that PARP-1 bound to the N-terminus of WRN and to the C-terminus containing the RecQ-conserved (RQC) domain. WRN bound to the N-terminus of PARP-1 containing DNA binding and BRCA1 C-terminal (BRCT) domains. We show that unmodified PARP-1 inhibited both WRN exonuclease and helicase activities, and to our knowledge is the only known WRN protein partner that inactivates both of the WRN's catalytic activities suggesting a biologically significant regulation. Moreover, this dual inhibition seems to be specific for PARP-1, as PARP-2 did not affect WRN helicase activity and only slightly inhibited WRN exonuclease activity. The differential effect of PARP-1 and PARP-2 on WRN catalytic activity was not due to differences in affinity for WRN or the DNA substrate. Finally, we demonstrate that the inhibition of WRN by PARP-1 was influenced by the poly(ADP-ribosyl)ation state of PARP-1. The biological relevance of the specific modulation of WRN catalytic activities by PARP-1 are discussed in the context of pathways in which these proteins may function together, namely in the repair of DNA strand breaks.
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PMID:Poly(ADP-ribose) polymerase 1 regulates both the exonuclease and helicase activities of the Werner syndrome protein. 1529 49

The Rothmund-Thomson syndrome (growth retardation, skin and bone defects, predisposition to cancer) and the RAPADILINO syndrome are caused by mutations in the RECQL4 gene. The 133 kDa RECQL4 is a putative DNA helicase, a member of the family that includes the BLM and WRN helicases. The latter are mutated, respectively, in the Bloom and Werner syndromes, whose manifestations include predisposition to cancer. Using antibodies to human RECQL4, we found that the bulk of RECQL4 was present in a cytoplasmic extract of HeLa cells, in contrast to the largely nuclear BLM and WRN helicases. However, in untransformed WI-38 fibroblasts, RECQL4 was found to be largely nuclear, and was present at significantly lower total levels than in transformed HeLa cells. RECQL4 from HeLa cells was isolated as a stable complex with UBR1 and UBR2. These 200 kDa proteins are ubiquitin ligases of the N-end rule pathway, whose substrates include proteins with destabilizing N-terminal residues. The functions of this proteolytic pathway include the regulation of peptide import, chromosome stability, meiosis, apoptosis and cardiovascular development. Although the known role of UBR1 and UBR2 is to mediate polyubiquitylation (and subsequent degradation) of their substrates, the UBR1/2-bound RECQL4 was not ubiquitylated in vivo, and was a long-lived protein in HeLa cells. The isolated RECQL4-UBR1/2 complex had a DNA-stimulated ATPase activity, but was inactive in DNA-based assays for helicases and translocases, the assays in which the BLM helicase was active. We discuss ramifications of these results, possible functions of RECQL4, and the involvement of the N-end rule pathway.
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PMID:RECQL4, mutated in the Rothmund-Thomson and RAPADILINO syndromes, interacts with ubiquitin ligases UBR1 and UBR2 of the N-end rule pathway. 1531 57

Human WRNIP1, a Werner DNA helicase interacting protein 1, was expressed in insect cells and E. coli. The purified protein behaved as a homo-oligomeric complex with a native molecular mass indicative of an octamer, and the complex copurified with an ATPase activity that was stimulated by double-stranded DNA ends. As suggested by genetic studies of budding yeast WRNIP1/Mgs1, the purified human WRNIP1 complex interacted physically with human DNA polymerase delta (pol delta), stimulating its DNA synthesis activity more than fivefold in the presence or absence of proliferating cell nuclear antigen. Analysis of reaction products demonstrated the stimulation to be partly due to an increased processivity of pol delta but more importantly to an increase in its initiation frequency. Addition of ATP to reactions partially suppressed stimulation by WRNIP1. Furthermore, a mutant WRNIP1 lacking ATPase activity could stimulate pol delta normally but was insensitive to suppression by ATP. These results indicate that WRNIP1 functions as a modulator for initiation or restart events during pol delta-mediated DNA synthesis and that its ATPase activity is utilized to sense DNA ends and to regulate the extent of stimulation.
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PMID:Human Werner helicase interacting protein 1 (WRNIP1) functions as a novel modulator for DNA polymerase delta. 1567 Feb 10

Nuclear DNA helicase II (NDH II), alternatively named RNA helicase A, is involved in transcription and RNA processing. Here, we report that NDH II interacts with the Werner syndrome helicase WRN, an enzyme associated with premature aging and predisposition to tumorigenesis. NDH II was co-purified with WRN, DNA polymerase delta, and replication protein A (70 kDa) during several steps of conventional column chromatography. Co-immunoprecipitations revealed an association between NDH II, WRN, and polymerase delta. We demonstrate a direct protein-protein interaction between WRN and NDH II that is mediated by the N-terminal double-strand RNA-binding domain II and C-terminal RGG box of NDH II and the N-terminal exonuclease domain of WRN. WRN inhibited the DNA-dependent NTPase and DNA helicase activities of NDH II. On the other hand, the 3' --> 5' exonuclease activity of WRN was increased by the presence of NDH II. NDH II directly stimulated the exonuclease domain of WRN, whereas the exonuclease domain of WRN suppressed the DNA-dependent (but not RNA-dependent) ATPase activity of NDH II. These results suggest that the double-strand RNA-binding domain II and RGG box of NDH II together form a protein-protein interaction surface that contacts the exonuclease domain of WRN. Furthermore, NDH II enhanced the degradation of D-loop DNA by the WRN exonuclease. Taken together, these results suggest that NDH II plays a role in promoting the DNA processing function of WRN, which in turn might be necessary for maintaining genomic stability.
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PMID:Nuclear DNA helicase II (RNA helicase A) interacts with Werner syndrome helicase and stimulates its exonuclease activity. 1599 49

Werner syndrome is a genetic disease characterized by early ageing, excess cancer risk, high incidence of type II diabetes mellitus, early atherosclerosis, ocular cataracts, and osteoporosis. The protein encoded by the defective gene, WRN (WRNp) associates with three activities, that is, a RecQ DNA helicase, 3'-5'-exonuclease and ATPase activities. By highlighting the DNA helicase activity, a widespread consensus in WS-associated defect(s) has been established, pointing toward a deficiency in maintaining DNA integrity. However, a possible involvement of redox pathways in WS may be suggested by several lines of evidence that include: (i) the multiple functions and interactions of WRNp with oxidative stress-related activities and factors; (ii) the pleiotropic WS clinical phenotype encompassing a number of oxidative stress-related pathologies; (iii) redox-related toxicity mechanisms of several xenobiotics exerting excess toxicity in WS cells; (iv) recent in vivo and in vitro findings of redox abnormalities in WS patients and in WS cells. The working hypothesis is raised that a deficiency in WRNp, and the pleiotropic clinical phenotype in WS patients may provide the basis to envision an underlying in vivo prooxidant state, which causes oxidative damage to biomolecules, with multiple oxidative stress-related alterations, resulting in multi-faceted clinical consequences.
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PMID:Multiple involvement of oxidative stress in Werner syndrome phenotype. 1633 57

Werner syndrome is a segmental progeroid disease characterized by increased cancer and acceleration of specific age-related phenotypes, due to loss of a protein known as WRN. Extensive research over the last decade has revealed much about WRN biochemistry and the etiology of Werner syndrome. WRN possesses multiple DNA-dependent enzymatic activities (ATPase, helicase, exonuclease, and strand annealing) and interacts with factors having established roles in DNA metabolic pathways. Although the exact functions of WRN remain unclear, accumulating evidence points to roles in proper resolution of replication blockage and in telomere maintenance. If WRN function is lost (as exemplified in cells from Werner patients), problems with replication and DNA damage processing arise, probably resulting in an increased number or persistence of strand breaks. In turn, these events lead to chromosomal and telomeric abnormalities or activate checkpoints that bring about early senescence or increased apoptosis. Thus, elevated cancer incidence associated with Werner syndrome is due to increased chromosomal changes, while the accelerated aging characteristics probably stem from telomere dysfunction leading to accumulation of non-functional senescent cells or excessive apoptotic cell death over time. More research is needed to determine whether these specific DNA-dependent mechanisms contribute to development of aging characteristics in normal individuals.
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PMID:Werner syndrome: molecular insights into the relationships between defective DNA metabolism, genomic instability, cancer and aging. 1672 Mar 42

Werner syndrome (WS) is a premature aging disorder characterized by genomic instability and increased cancer risk (Martin, 1978). The WRN gene product defective in WS belongs to the RecQ family of DNA helicases (Yu et al., 1996). Mutations in RecQ family members BLM and RecQ4 result in two other disorders associated with elevated chromosomal instability and cancer, Bloom syndrome and Rothmund-Thomson syndrome, respectively (for review see Opresko et al., 2004a). RecQ helicase mutants display defects in DNA replication, recombination, and repair, suggesting a role for RecQ helicases in maintaining genomic integrity. The WRN gene encodes a 1,432 amino acid protein that has several catalytic activities (Brosh and Bohr, 2002) (Fig. 1). WRN is a DNA-dependent ATPase and utilizes the energy from ATP hydrolysis to unwind double-stranded DNA. WRN is also a 3' to 5' exonuclease, consistent with the presence of three conserved exonuclease motifs homologous to the exonuclease domain of Escherichia coli DNA polymerase I and RNase D. Most recently, WRN (Machwe et al., 2005) and other human RecQ helicases (Garcia et al., 2004; Machwe et al., 2005; Sharma et al., 2005) have been reported to possess an intrinsic single-strand annealing activity. In addition to its catalytic activities, WRN interacts with a number of proteins involved in various aspects of DNA metabolism. To understand the role of WRN in the maintenance of genome stability, a number of laboratories have undertaken a thorough characterization of its molecular and cellular functions. Here, we describe methods and approaches used for the functional and mechanistic analysis of WRN helicase or exonuclease activity. Protocols for measuring ATP hydrolysis, DNA binding, and catalytic unwinding or exonuclease activity of WRN protein are provided. Application of these procedures should enable the researcher to address fundamental questions regarding the biochemical properties of WRN or related helicases or nucleases, which would serve as a platform for further investigation of its molecular and cellular functions.
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PMID:Enzymatic mechanism of the WRN helicase/nuclease. 1679 95

Werner syndrome (WS) is an autosomal recessive progeroid disease characterized by genomic instability. WRN gene encodes one of the RecQ helicase family proteins, WRN, which has ATPase, helicase, exonuclease and single stranded DNA annealing activities. There is accumulating evidence suggesting that WRN contributes to the maintenance of genomic integrity through its involvement in DNA repair, replication and recombination. The role of WRN in these pathways can be modulated by its post-translational modifications in response to DNA damage. Here, we review the functional consequences of post-translational modifications on WRN as well as specific DNA repair pathways where WRN is involved and discuss how these modifications affect DNA repair pathways.
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PMID:The role of WRN in DNA repair is affected by post-translational modifications. 1711 23

The premature aging and cancer-prone disease Werner syndrome stems from loss of WRN protein function. WRN deficiency causes replication abnormalities, sensitivity to certain genotoxic agents, genomic instability and early replicative senescence in primary fibroblasts. As a RecQ helicase family member, WRN is a DNA-dependent ATPase and unwinding enzyme, but also possesses strand annealing and exonuclease activities. RecQ helicases are postulated to participate in pathways responding to replication blockage, pathways possibly initiated by fork regression. In this study, a series of model replication fork substrates were used to examine the fork regression capability of WRN. Our results demonstrate that WRN catalyzes fork regression and Holliday junction formation. This process is an ATP-dependent reaction that is particularly efficient on forks containing single-stranded gaps of at least 11-13 nt on the leading arm at the fork junction. Importantly, WRN exonuclease activity, by digesting the leading daughter strand, enhances regression of forks with smaller gaps on the leading arm, thus creating an optimal structure for regression. Our results suggest that the multiple activities of WRN cooperate to promote replication fork regression. These findings, along with the established cellular consequences of WRN deficiency, strongly support a role for WRN in regression of blocked replication forks.
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PMID:Replication fork regression in vitro by the Werner syndrome protein (WRN): holliday junction formation, the effect of leading arm structure and a potential role for WRN exonuclease activity. 1771 3

BLM, the protein mutated in Bloom's syndrome, possesses a helicase activity that can dissociate DNA structures, including the Holliday junction, expected to arise during homologous recombination. BLM is stably associated with topoisomerase IIIalpha (Topo IIIalpha) and the BLAP75 protein. The BLM-Topo IIIalpha-BLAP75 (BTB) complex can efficiently resolve a DNA substrate that harbors two Holliday junctions (the double Holliday junction) in a non-crossover manner. Here we show that the Holliday junction unwinding activity of BLM is greatly enhanced as a result of its association with Topo IIIalpha and BLAP75. Enhancement of this BLM activity requires both Topo IIIalpha and BLAP75. Importantly, Topo IIIalpha cannot be substituted by Escherichia coli Top3, and the Holliday junction unwinding activity of BLM-related helicases WRN and RecQ is likewise impervious to Topo IIIalpha and BLAP75. However, the topoisomerase activity of Topo IIIalpha is dispensable for the enhancement of the DNA unwinding reaction. We have also ascertained the requirement for the BLM ATPase activity in double Holliday junction dissolution and DNA unwinding by constructing, purifying, and characterizing specific mutant variants that lack this activity. These results provide valuable information concerning how the functional integrity of the BTB complex is governed by specific protein-protein interactions among the components of this complex and the enzymatic activities of BLM and Topo IIIalpha.
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PMID:Holliday junction processing activity of the BLM-Topo IIIalpha-BLAP75 complex. 1772 55


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