Single-strand DNA Binding by the Helix-Hairpin-Helix Domain of XPF Contributes to Substrate Specificity of ERCC1-XPF

The nucleotide excision repair protein complex ERCC1-XPF is required for incision of DNA upstream of DNA damage. Functional studies have provided insights into the binding of ERCC1-XPF to various DNA substrates. However, because no structure for the ERCC1-XPF-DNA complex has been determined, the mechanism of substrate recognition remains elusive. Here we biochemically characterize the substrate preferences of the helix-hairpin-helix (HhH) domains of XPF and ERCC-XPF and show that the binding to single-stranded DNA (ssDNA)/dsDNA junctions is dependent on joint binding to the DNA binding domain of ERCC1 and XPF. We reveal that the homodimeric XPF is able to bind various ssDNA sequences but with a clear preference for guanine-containing substrates. NMR titration experiments and in vitro DNA binding assays also show that, within the heterodimeric ERCC1-XPF complex, XPF specifically recognizes ssDNA. On the other hand, the HhH domain of ERCC1 preferentially binds dsDNA through the hairpin region. The two separate non-overlapping DNA binding domains in the ERCC1-XPF heterodimer jointly bind to an ssDNA/dsDNA substrate and, thereby, at least partially dictate the incision position during damage removal. Based on structural models, NMR titrations, DNA-binding studies, site-directed mutagenesis, charge distribution, and sequence conservation, we propose that the HhH domain of ERCC1 binds to dsDNA upstream of the damage, and XPF binds to the non-damaged strand within a repair bubble

Алгоритм проведення попередньої дегазації методом гравітаційного знещільнення вуглепородного масиву

Приводится алгоритм проведения предварительной дегазации при применении нового способа гравитационной разгрузки на угольных месторождениях Донбасса и Львовско-Волынского бассейнов. Среди главных эффектов, которые будут получены при внедрении способа – обеспечение безопасности работ на шахтах и промышленная добыча метана.The algorithm of realization of pre-degassing with using new way of gravitational unloading on coal fields of Donbas and Lviv-Volyn basins is given in this article. Guaranteeing of safety works in mines and industrial methane-mining are between main effects which will be got on conditions of application of this method

Structural and biochemical characterization of NarE, an iron-containing ADP-ribosyltransferase from Neisseria meningitidis.

Abstract NarE is a 16 kDa protein identified from Neisseria meningitidis, one of the bacterial pathogens responsible for meningitis. NarE belongs to the family of ADP-ribosyltransferases (ADPRT) and catalyzes the transfer of ADP-ribose moieties to arginine residues in target protein acceptors. Many pathogenic bacteria utilize ADP-ribosylating toxins to modify and alter essential functions of eukaryotic cells. NarE is further the first ADPRT which could be shown to bind iron through a Fe-S center, which is crucial for the catalytic activity. Here we present the NMR solution structure of NarE, which shows structural homology to other ADPRTs. Using NMR titration experiments we could identify from Chemical Shift Perturbation data both the NAD binding site, which is in perfect agreement with a consensus sequence analysis between different ADPRTs, as well as the iron coordination site, which consists of 2 cysteines and 2 histidines. This atypical iron coordination is also capable to bind zinc. These results could be fortified by site-directed mutagenesis of the catalytic region, which identified two functionally crucial residues. We could further identify a main interaction region of NarE with antibodies using two complementary methods based on antibody immobilization, proteolytic digestion, and mass spectrometry. This study combines structural and functional features of NarE providing for the first time a characterization of an iron-dependent ADPRT

The Subak in Diaspora: Balinese Farmers and the Subak in South Sulawesi

The subak has a long history as an irrigators’ institution on Bali. It has also spread across Indonesia along with Balinese farmers who were resettled by colonial and post-colonial governments or who have migrated spontaneously since colonial times. While subaks have been much researched in Bali itself, little is known about subaks outside Bali. Luwu District in South Sulawesi is one of the areas where thousands of Balinese families settled in the last four decades. Based on research in this transmigration area, this paper analyzes the emergence and development of the subak in relation to the development of irrigation infrastructure of a state-built irrigation system. A comparison between two Balinese settlements in the same system shows that differences in infrastructural and managerial conditions and arrangements between parts of the irrigation system were major determinants of the institutional space allowed for the subak and ways in which the subaks developed