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single-stranded DNA binding protein

Isidoro Feliciello, Davor Zahradka, Ksenija Zahradka, Siniša Ivanković, Nikolina Puc, Damir Đermić
Double strand breaks (DSBs) in E. coli chromosome (such as those induced by gamma rays) are repaired by recombination repair, during which a certain amount of DNA gets degraded. We monitored DNA degradation in gamma-irradiated cells to assess processing of DSBs. DNA degradation in irradiated cells is regulated by RecA protein concentration and its affinity of ssDNA binding, as well as by exonucleases that trim 3'-terminated ss tails. Here we determined the effects of proteins that affect formation and stability of RecA nucleofilaments on DNA degradation and cell survival...
March 16, 2018: Biochimie
Daniela Cecconi, Luca Dalle Carbonare, Antonio Mori, Samuele Cheri, Michela Deiana, Jessica Brandi, Vincenzo Degaetano, Valentina Masiero, Giulio Innamorati, Monica Mottes, Giovanni Malerba, Maria Teresa Valenti
Melanoma is an aggressive skin cancer; an early detection of the primary tumor may improve its prognosis. Despite many genes have been shown to be involved in melanoma, the full framework of melanoma transformation has not been completely explored. The characterization of pathways involved in tumor restraint in in vitro models may help to identify oncotarget genes. We therefore aimed to probe novel oncotargets through an integrated approach involving proteomic, gene expression and bioinformatic analysis We investigated molecular modulations in melanoma cells treated with ascorbic acid, which is known to inhibit cancer growth at high concentrations...
February 20, 2018: Oncotarget
Xiubao Chang, Yuexian Hou
Genome editing is a powerful tool to modify a specific gene and to correct a disease-causing mutation. Recently developed new techniques, such as zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALEN) and clustered regularly interspaced short palindromic repeats/Cas9 (CRISPR/Cas9), significantly facilitate the progression in this field. However, mutations associated with the double strand DNA breaks (DSBs) introduced by these systems hampered their direct usage in clinic. In order to prevent the mutations caused by DSBs, we have designed a novel mean to induce homology-directed recombination (HDR) without DSBs, i...
2018: International Journal of Biochemistry and Molecular Biology
Jia Zhou, Jiaqi Li, Rodolfo B Serafim, Steven Ketchum, Catarina G Ferreira, Jessica C Liu, Kathryn A Coe, Brendan D Price, Timur Yusufzai
CHD1 is a conserved chromatin remodeling enzyme required for development and linked to prostate cancer in adults, yet its role in human cells is poorly understood. Here, we show that targeted disruption of the CHD1 gene in human cells leads to a defect in early double-strand break (DSB) repair via homologous recombination (HR), resulting in hypersensitivity to ionizing radiation as well as PARP and PTEN inhibition. CHD1 knockout cells show reduced H2AX phosphorylation (γH2AX) and foci formation as well as impairments in CtIP recruitment to the damaged sites...
February 26, 2018: Nucleic Acids Research
Amandeep Singh, M Vijayan, Umesh Varshney
In addition to the canonical Single Stranded DNA Binding (SSBa) protein, many bacterial species, including mycobacteria, have a paralogous SSBb. The SSBb proteins have not been well characterized. While in B. subtilis, SSBb has been shown to be involved in genetic recombination; in S. coelicolor it mediates chromosomal segregation during sporulation. Sequence analysis of SSBs from mycobacterial species suggests low conservation of SSBb proteins, as compared to the conservation of SSBa proteins. Like most bacterial SSB proteins, M...
January 2018: Tuberculosis
Quinn Li, Laura Folly da Silva Constantino, M Ashley Spies
Discovery of novel tool compounds and drug leads against a range of unorthodox protein targets has pushed both experimental screening methodologies as well as the field of structure-based design to the limit in recent years. Increasingly, it has been recognized that some of the most desirable targets for the development of small-molecule effectors are actually protein-protein and protein-nucleic acid interactions. There are numerous nontrivial challenges to pursuing small-molecule lead compounds directed toward PPIs and PNIs: relatively shallow cavities, large surface areas that are natively complexed to macromolecules, complex patterns of interstitial waters, a paucity of "hot spots," large conformational changes upon ligand binding, etc...
2018: Methods in Enzymology
Nidhi Sharma, Srinivas Chakravarthy, Matthew J Longley, William C Copeland, Aishwarya Prakash
The 16.5 kb mitochondrial genome is subjected to damage from reactive oxygen species (ROS) generated in the cell during normal cellular metabolism and external sources such as ionizing radiation and ultraviolet light. ROS cause harmful damage to DNA bases that could result in mutagenesis and various diseases, if not properly repaired. The base excision repair (BER) pathway is the primary pathway involved in maintaining the integrity of mtDNA. Several enzymes that partake in BER within the nucleus have also been identified in the mitochondria...
March 6, 2018: DNA Repair
Hongjian Gong, Yuxi Zhang, Kunpeng Jiang, Shengfan Ye, Shuming Chen, Qinghe Zhang, Jinrong Peng, Jun Chen
Tumour repressor p53 isoform Δ133p53 is a target gene of p53 and an antagonist of p53-mediated apoptotic activity. We recently demonstrated that Δ133p53 promotes DNA double-strand break (DSB) repair by upregulating transcription of the repair genes RAD51, LIG4 and RAD52 in a p53-independent manner. However, Δ133p53 lacks the transactivation domain of full-length p53, and the mechanism by which it exerts transcriptional activity independently of full-length p53 remains unclear. In this report, we describe the accumulation of high levels of both Δ133p53 and p73 (a p53 family member) at 24 h post γ-irradiation (hpi)...
March 6, 2018: Cell Death and Differentiation
Maja Sidstedt, Johannes Hedman, Erica L Romsos, Leticia Waitara, Lars Wadsö, Carolyn R Steffen, Peter M Vallone, Peter Rådström
Blood samples are widely used for PCR-based DNA analysis in fields such as diagnosis of infectious diseases, cancer diagnostics, and forensic genetics. In this study, the mechanisms behind blood-induced PCR inhibition were evaluated by use of whole blood as well as known PCR-inhibitory molecules in both digital PCR and real-time PCR. Also, electrophoretic mobility shift assay was applied to investigate interactions between inhibitory proteins and DNA, and isothermal titration calorimetry was used to directly measure effects on DNA polymerase activity...
March 5, 2018: Analytical and Bioanalytical Chemistry
Yinhua Zhang, Nathan A Tanner
A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.
March 2, 2018: Scientific Reports
Tanja Narancic, Elisa Scollica, Gerard Cagney, Kevin E O'Connor
Polyhydroxybutyrate (PHB), a biodegradable polymer accumulated by bacteria is deposited intracellularly in the form of inclusion bodies often called granules. The granules are supramolecular complexes harbouring a varied number of proteins on their surface, which have specific but incompletely characterised functions. By comparison with other organisms that produce biodegradable polymers, only two phasins have been described to date for Rhodosprillum rubrum, raising the possibility that more await discovery...
March 1, 2018: Microbiology
Tongbo Wu, Yufei Yang, Wei Chen, Jiayu Wang, Ziyu Yang, Shenlin Wang, Xianjin Xiao, Mengyuan Li, Meiping Zhao
Lambda exonuclease (λ exo) plays an important role in the resection of DNA ends for DNA repair. Currently, it is also a widely used enzymatic tool in genetic engineering, DNA-binding protein mapping, nanopore sequencing and biosensing. Herein, we disclose two noncanonical properties of this enzyme and suggest a previously undescribed hydrophobic interaction model between λ exo and DNA substrates. We demonstrate that the length of the free portion of the substrate strand in the dsDNA plays an essential role in the initiation of digestion reactions by λ exo...
February 27, 2018: Nucleic Acids Research
E E Alemasova, K N Naumenko, N A Moor, O I Lavrik
Apurinic/apyrimidinic (AP) sites are among the most frequent DNA lesions. The first step in the AP site repair involves the magnesium-dependent enzyme AP endonuclease 1 (APE1) that catalyzes hydrolytic cleavage of the DNA phosphodiester bond at the 5' side of the AP site, thereby generating a single-strand DNA break flanked by the 3'-OH and 5'-deoxyribose phosphate (dRP) groups. Increased APE1 activity in cancer cells might correlate with tumor chemoresistance to DNA-damaging treatment. It has been previously shown that the multifunctional oncoprotein Y-box-binding protein 1 (YB-1) interacts with APE1 and inhibits APE1-catalyzed hydrolysis of AP sites in single-stranded DNAs...
December 2017: Biochemistry. Biokhimii︠a︡
Yuan Wang, Weihang Chai
Coats plus syndrome is a complex genetic disorder that can be caused by mutations in genes encoding the CTC1-STN1-TEN1 (CST) complex, a conserved single-stranded DNA binding protein complex. Studies have demonstrated that mutations identified in Coats plus patients are defective in telomere maintenance, and concluded that Coats plus may be caused by telomere dysfunction. Recent studies have established that CST also plays an important role in countering replication stress and protecting the stability of genomic fragile sites...
February 22, 2018: Nucleic Acids Research
Olga A Kladova, Milena Bazlekowa-Karaban, Sonia Baconnais, Olivier Piétrement, Alexander A Ishchenko, Bakhyt T Matkarimov, Danila A Iakovlev, Andrey Vasenko, Olga S Fedorova, Eric Le Cam, Barbara Tudek, Nikita A Kuznetsov, Murat Saparbaev
The base excision repair (BER) pathway consists of sequential action of DNA glycosylase and apurinic/apyrimidinic (AP) endonuclease necessary to remove a damaged base and generate a single-strand break in duplex DNA. Human multifunctional AP endonuclease 1 (APE1, a.k.a. APEX1, HAP-1, or Ref-1) plays essential roles in BER by acting downstream of DNA glycosylases to incise a DNA duplex at AP sites and remove 3'-blocking sugar moieties at DNA strand breaks. Human 8-oxoguanine-DNA glycosylase (OGG1), methyl-CpG-binding domain 4 (MBD4, a...
February 11, 2018: DNA Repair
Shankar Shastry, Olga Steinberg-Neifach, Neal Lue, Michael D Stone
Telomerase is a specialized enzyme that maintains telomere length by adding DNA repeats to chromosome ends. The catalytic protein subunit of telomerase utilizes the integral telomerase RNA to direct telomere DNA synthesis. The telomerase essential N-terminal (TEN) domain is required for enzyme function; however, the precise mechanism of the TEN domain during catalysis is not known. We report a single-molecule study of dynamic TEN-induced conformational changes in its nucleic acid substrates. The TEN domain from the yeast Candida parapsilosis (Cp) exhibits a strong binding preference for double-stranded nucleic acids, with particularly high affinity for an RNA-DNA hybrid mimicking the template-product complex...
February 21, 2018: Nucleic Acids Research
Graeme A King, Andreas S Biebricher, Iddo Heller, Erwin Peterman, Gijs Wuite
The ability to measure mechanics and forces in biological nanostructures, such as DNA, proteins and cells, is of great importance as a means to analyze biomolecular systems. However, current force detection methods often require specialized instrumentation. Here, we present a novel and versatile method to quantify tension in molecular systems locally and in real time using intercalated DNA fluorescence. This approach can report forces over a range of ~1-65 pN with a resolution of 1-3 pN, using commercially available intercalating dyes and a general-purpose fluorescence microscope...
February 23, 2018: Nano Letters
Zorik Chilingaryan, Stephen J Headey, Allen T Y Lo, Zhi-Qiang Xu, Gottfried Otting, Nicholas E Dixon, Martin J Scanlon, Aaron J Oakley
In bacteria, the DnaG primase is responsible for synthesis of short RNA primers used to initiate chain extension by replicative DNA polymerase(s) during chromosomal replication. Among the proteins with which Escherichia coli DnaG interacts is the single-stranded DNA-binding protein, SSB. The C-terminal hexapeptide motif of SSB (DDDIPF; SSB-Ct) is highly conserved and is known to engage in essential interactions with many proteins in nucleic acid metabolism, including primase. Here, fragment-based screening by saturation-transfer difference nuclear magnetic resonance (STD-NMR) and surface plasmon resonance assays identified inhibitors of the primase/SSB-Ct interaction...
February 22, 2018: Antibiotics
Liisa T Chisty, Daniela Quaglia, Martin R Webb
Single-stranded DNA (ssDNA) is a product of many cellular processes that involve double-stranded DNA, for example during DNA replication and repair, and is formed transiently in many others. Measurement of ssDNA formation is fundamental for understanding many such processes. The availability of a fluorescent biosensor for the determination of single-stranded DNA provides an important route to achieve this. Single-stranded DNA binding proteins (SSBs) protect ssDNA from degradation, but can be displaced to allow processing of the ssDNA...
2018: PloS One
Olivia Yang, Taekjip Ha
Single-stranded DNA-binding protein (SSB) is important not only for the protection of single-stranded DNA (ssDNA) but also for the recruitment of other proteins for DNA replication, recombination, and repair. The interaction of SSB with ssDNA is highly dynamic as it exists as an intermediate during cellular processes that unwind dsDNA. It has been proposed that SSB redistributes itself among multiple ssDNA segments, but transient intermediates are difficult to observe in bulk experiments. We can use single-molecule FRET microscopy to observe intermediates of the transfer of a single Escherichia coli SSB from one ssDNA strand to another or exchange of one SSB for another on a single ssDNA in real time...
2018: Methods in Enzymology
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