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Helicase Domain (helicase + domain)
Selected AbstractsRole of DNA polymerase , in tolerance of endogenous and exogenous DNA damage in mouse B cellsGENES TO CELLS, Issue 2 2006Akiko Ukai DNA polymerase , (Pol,) is a family A polymerase that contains an intrinsic helicase domain. To investigate the function of Pol, in mammalian cells, we have inactivated its polymerase activity in CH12 mouse B lymphoma cells by targeted deletion of the polymerase core domain that contains the catalytic aspartic acid residue. Compared to parental CH12 cells, mutant cells devoid of Pol, polymerase activity exhibited a slightly reduced growth rate, accompanied by increased spontaneous cell death. In addition, mutant cells showed elevated sensitivity to mitomycin C, cisplatin, etoposide, ,-irradiation and ultraviolet (UV) radiation. Interestingly, mutant cells were more sensitive to the alkylating agent methyl methanesulfonate (MMS) than parental cells. This elevated MMS sensitivity relative to WT cells persisted in the presence of methoxyamine, an inhibitor of the major base excision repair (BER) pathway, suggesting that Pol, is involved in tolerance of MMS through a mechanism that appears to be different from BER. These results reveal an important role for Pol, in preventing spontaneous cell death and in tolerance of not only DNA interstrand cross-links and double strand breaks but also UV adducts and alkylation damage in mammalian lymphocytes. [source] Sequence similarities between Raspberry leaf mottle virus, Raspberry leaf spot virus and the closterovirus Raspberry mottle virusANNALS OF APPLIED BIOLOGY, Issue 3 2010W.J. McGavin A sequencing study was performed to determine the relationship between Raspberry mottle virus (RMoV), a newly identified tentative closterovirus found in the United States, and Raspberry leaf mottle virus (RLMV) and Raspberry leaf spot virus (RLSV), which have been known for many years to be components of Raspberry mosaic disease (RMD) in the UK and Europe but which have not been characterised at the molecular level. Cloning and sequencing of cDNAs amplified by reverse transcription-PCR revealed the presence of closteroviruses with high sequence similarity to RMoV in infected plants from the SCRI Rubus virus collection, as well as in a number of samples collected from RMD-symptomatic raspberry plants located at different farms in Scotland and England. These results suggest that RMoV, RLMV and RLSV are isolates of the same virus and we propose that they all should be referred to as RLMV, which was the first of these viruses to be described. Many of the field samples were also infected with a second closterovirus isolate, parts of which could be amplified using RLMV-derived primers. The coat protein amino acid sequences of RLMV and the second virus (PM1) were only 78% identical, even though helicase domain and RNA-dependent RNA polymerase (RDRP) domain sequences were more than 97% identical between RLMV and PM1. [source] Mutations in PHD-like domain of the ATRX gene correlate with severe psychomotor impairment and severe urogenital abnormalities in patients with ATRX syndromeCLINICAL GENETICS, Issue 1 2006C Badens Mutations in ATRX are associated with a wide and clinically heterogeneous spectrum of X-linked mental retardation syndromes. The ATRX protein, involved in chromatin remodelling, belongs to the family of SWI/SNF DNA helicases and contains a plant homeodomain (PHD)-like domain. To date, more than 60 different mutations have been reported in ATRX. One of them is recurrent and accounts for 20% of all the reported mutations, whereas all others are private. Most mutations are clustered in the two major functional domains, the helicase and the PHD-like domain. So far, no clear genotype,phenotype correlation has been established, with exception to the rare truncating mutations located at the C-terminal part of the protein, which are consistently associated with severe urogenital defects. In this study, we report the molecular analysis performed in 16 families positive for ATRX. Our findings indicate that, in addition to the previously described mutation ,hotspot' in the PHD-like domain, two other protein sections emerge as minor ,hotspots' in the helicase region encoded by exons 18,20 and 26,29, respectively, gathering 33% of all described mutations. Additionally, based on the clinical data collected for 22 patients from the 16 families, we observe that mutations in the PHD-like domain produce severe and permanent psychomotor deficiency, usually preventing patients from walking, as well as constant urogenital abnormalities, while mutations in the helicase domain lead to delayed but correct psychomotor acquisitions together with mild or absent urogenital abnormalities. In summary, mutations in the helicase domain are associated with milder phenotypes than mutations in the PHD-like domain. [source] Authentic interdomain communication in an RNA helicase reconstituted by expressed protein ligation of two helicase domainsFEBS JOURNAL, Issue 2 2007Anne R. Karow RNA helicases mediate structural rearrangements of RNA or RNA,protein complexes at the expense of ATP hydrolysis. Members of the DEAD box helicase family consist of two flexibly connected helicase domains. They share nine conserved sequence motifs that are involved in nucleotide binding and hydrolysis, RNA binding, and helicase activity. Most of these motifs line the cleft between the two helicase domains, and extensive communication between them is required for RNA unwinding. The two helicase domains of the Bacillus subtilis RNA helicase YxiN were produced separately as intein fusions, and a functional RNA helicase was generated by expressed protein ligation. The ligated helicase binds adenine nucleotides with very similar affinities to the wild-type protein. Importantly, its intrinsically low ATPase activity is stimulated by RNA, and the Michaelis,Menten parameters are similar to those of the wild-type. Finally, ligated YxiN unwinds a minimal RNA substrate to an extent comparable to that of the wild-type helicase, confirming authentic interdomain communication. [source] Isolation and characterization of the RAD54 gene from Arabidopsis thalianaTHE PLANT JOURNAL, Issue 6 2006Keishi Osakabe Summary Homologous recombination (HR) is an essential process in maintaining genome integrity and variability. In eukaryotes, the Rad52 epistasis group proteins are involved in meiotic recombination and/or HR repair. One member of this group, Rad54, belongs to the SWI2/SNF2 family of DNA-stimulated ATPases. Recent studies indicate that Rad54 has important functions in HR, both as a chromatin remodelling factor and as a mediator of the Rad51 nucleoprotein filament. Despite the importance of Rad54 in HR, no study of Rad54 from plants has yet been performed. Here, we cloned the full-length AtRAD54 cDNA sequence; an open reading frame of 910 amino acids encodes a protein with a predicted molecular mass of 101.9 kDa. Western blotting analysis showed that the AtRad54 protein was indeed expressed as a protein of approximately 110 kDa in Arabidopsis. The predicted protein sequence of AtRAD54 contains seven helicase domains, which are conserved in all other Rad54s. Yeast two-hybrid analysis revealed an interaction between Arabidopsis Rad51 and Rad54. AtRAD54 transcripts were found in all tissues examined, with the highest levels of expression in flower buds. Expression of AtRAD54 was induced by , -irradiation. A T-DNA insertion mutant of AtRAD54 devoid of full-length AtRAD54 expression was viable and fertile; however, it showed increased sensitivity to , -irradiation and the cross-linking reagent cisplatin. In addition, the efficiency of somatic HR in the mutant plants was reduced relative to that in wild-type plants. Our findings point to an important role for Rad54 in HR repair in higher plants. [source] | ||||||||||