Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.6.1.3 (ATPase)
 65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Xeroderma pigmentosum (XP), a genetically heterogeneous human disease, results from a defect in nucleotide excision repair of ultraviolet-damaged DNA. XP patients are extremely sensitive to sunlight and suffer from a high incidence of skin cancers. Cell fusion studies have identified seven XP complementation groups, A-G. Group D is of particular interest as mutations in this gene can also cause Cockayne's syndrome and trichothiodystrophy. The XPD gene was initially named ERCC2 (excision repair cross complementing) as it was cloned using human DNA to complement the ultraviolet sensitivity of a rodent cell line. We have purified the XPD protein to near homogeneity and show that it possesses single-stranded DNA-dependent ATPase and DNA helicase activities. We tested whether XPD can substitute for its yeast counterpart RAD3, which is essential for excision repair and for cell viability. Expression of the XPD gene in Saccharomyces cerevisiae can complement the lethality defect of a mutation in the RAD3 gene, suggesting that XPD is an essential gene in humans.
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PMID:Human xeroderma pigmentosum group D gene encodes a DNA helicase. 841 72

TFIIH is a high molecular weight complex with a remarkable dual function in nucleotide excision repair and initiation of RNA polymerase II transcription. Mutations in the largest subunits, the XPB and XPD helicases, are associated with three inherited disorders: xeroderma pigmentosum, Cockayne's syndrome, and trichothiodystrophy. To facilitate the purification and biochemical characterization of this intricate complex, we generated a cell line stably expressing tagged XPB, allowing the immunopurification of the XPB protein and associated factors. Addition of two tags, a N-terminal hexameric histidine stretch and a C-terminal hemagglutinin epitope, to this highly conserved protein did not interfere with its functioning in repair and transcription. The hemagglutinin epitope allowed efficient TFIIH immunopurification to homogeneity from a fractionated whole cell extract in essentially one step. We conclude that the predominant active form of TFIIH is composed of nine subunits and that there is one molecule of XPB per TFIIH complex. The affinity-purified complex exhibits all expected TFIIH activities: DNA-dependent ATPase, helicase, C-terminal domain kinase, and participation in in vitro and in vivo nucleotide excision repair and in vitro transcription. The affinity purification procedure described here is fast and simple, does not require extensive chromatographic procedures, and yields highly purified, active TFIIH.
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PMID:Affinity purification of human DNA repair/transcription factor TFIIH using epitope-tagged xeroderma pigmentosum B protein. 942 74

TFIIH is a multisubunit protein complex involved in RNA polymerase II transcription and nucleotide excision repair, which removes a wide variety of DNA lesions including UV-induced photoproducts. Mutations in the DNA-dependent ATPase/helicase subunits of TFIIH, XPB and XPD, are associated with three inherited syndromes as follows: xeroderma pigmentosum with or without Cockayne syndrome and trichothiodystrophy. By using epitope-tagged XPD we purified mammalian TFIIH carrying a wild type or an active-site mutant XPD subunit. Contrary to XPB, XPD helicase activity was dispensable for in vitro transcription, catalytic formation of trinucleotide transcripts, and promoter opening. Moreover, in contrast to XPB, microinjection of mutant XPD cDNA did not interfere with in vivo transcription. These data show directly that XPD activity is not required for transcription. However, during DNA repair, neither 5' nor 3' incisions in defined positions around a DNA adduct were detected in the presence of TFIIH containing inactive XPD, although substantial damage-dependent DNA synthesis was induced by the presence of mutant XPD both in cells and cell extracts. The aberrant damage-dependent DNA synthesis caused by the mutant XPD does not lead to effective repair, consistent with the discrepancy between repair synthesis and survival in cells from a number of XP-D patients.
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PMID:TFIIH with inactive XPD helicase functions in transcription initiation but is defective in DNA repair. 1066 May 93

The transcription and DNA repair factor TFIIH is composed of 10 subunits. Mutations in the XPB, XPD, and p8 subunits are genetically linked to human diseases, including cancer. However, no reports of mutations in other TFIIH subunits have been reported in higher eukaryotes. Here, we analyze at genetic, molecular, and biochemical levels the Drosophila melanogaster p52 (DMP52) subunit of TFIIH. We found that DMP52 is encoded by the gene marionette in Drosophila and that a defective DMP52 produces UV light-sensitive flies and specific phenotypes during development: organisms are smaller than their wild-type siblings and present tumors and chromosomal instability. The human homologue of DMP52 partially rescues some of these phenotypes. Some of the defects observed in the fly caused by mutations in DMP52 generate trichothiodystrophy and cancer-like phenotypes. Biochemical analysis of DMP52 point mutations introduced in human p52 at positions homologous to those of defects in DMP52 destabilize the interaction between p52 and XPB, another TFIIH subunit, thus compromising the assembly of the complex. This study significantly extends the role of p52 in regulating XPB ATPase activity and, consequently, both its transcriptional and nucleotide excision repair functions.
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PMID:DNA repair and transcriptional deficiencies caused by mutations in the Drosophila p52 subunit of TFIIH generate developmental defects and chromosome fragility. 1733 30

Helicases have important roles in nucleic acid metabolism, and their prominence is marked by the discovery of genetic disorders arising from disease-causing mutations. Missense mutations can yield unique insight to molecular functions and basis for disease pathology. XPB or XPD missense mutations lead to Xeroderma pigmentosum, Cockayne's syndrome, Trichothiodystrophy, or COFS syndrome, suggesting that DNA repair and transcription defects are responsible for clinical heterogeneity. Complex phenotypes are also observed for RECQL4 helicase mutations responsible for Rothmund-Thomson syndrome, Baller-Gerold syndrome, or RAPADILINO. Bloom's syndrome causing missense mutations are found in the conserved helicase and RecQ C-terminal domain of BLM that interfere with helicase function. Although rare, patient-derived missense mutations in the exonuclease or helicase domain of Werner syndrome protein exist. Characterization of WRN separation-of-function mutants may provide insight to catalytic requirements for suppression of phenotypes associated with the premature aging disorder. Characterized FANCJ missense mutations associated with breast cancer or Fanconi anemia interfere with FANCJ helicase activity required for DNA repair and the replication stress response. For example, a FA patient-derived mutation in the FANCJ Iron-Sulfur domain was shown to uncouple its ATPase and translocase activity from DNA unwinding. Mutations in DDX11 (ChlR1) are responsible for Warsaw Breakage syndrome, a recently discovered autosomal recessive cohesinopathy. Ongoing and future studies will address clinically relevant helicase mutations and polymorphisms, including those that interfere with key protein interactions or exert dominant negative phenotypes (e.g., certain mutant alleles of Twinkle mitochondrial DNA helicase). Chemical rescue may be an approach to restore helicase activity in loss-of-function helicase disorders. Genetic and biochemical analyses of disease-causing missense mutations in human helicase disorders have led to new insights to the molecular defects underlying aberrant cellular and clinical phenotypes.
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PMID:Disease-causing missense mutations in human DNA helicase disorders. 2327 57

The eukaryotic XPD helicase is an essential subunit of TFIIH involved in both transcription and nucleotide excision repair (NER). Mutations in human XPD are associated with several inherited diseases such as xeroderma pigmentosum, Cockayne syndrome, and trichothiodystrophy. We performed a comparative analysis of XPD from Homo sapiens and Chaetomium thermophilum (a closely related thermostable fungal orthologue) to decipher the different molecular prerequisites necessary for either transcription or DNA repair. In vitro and in vivo assays demonstrate that mutations in the 4Fe4S cluster domain of XPD abrogate the NER function of TFIIH and do not affect its transcriptional activity. We show that the p44-dependent activation of XPD is promoted by the stimulation of its ATPase activity. Furthermore, we clearly demonstrate that XPD requires DNA binding, ATPase, and helicase activity to function in NER. In contrast, these enzymatic properties are dispensable for transcription initiation. XPD helicase is thus exclusively devoted to NER and merely acts as a structural scaffold to maintain TFIIH integrity during transcription.
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PMID:In TFIIH, XPD helicase is exclusively devoted to DNA repair. 2526 80