Diachronous Settlement on the Territory of the Turgay Trough

The article introduces into the scholarship the collection of findings of prehistoric material culture obtained at Bestamak during the stationary studies of 2001 field season. The total area of the archaeological site is 260,000 m2. It was excavated by Turgay expedition in the 1980s. Bestamak settlement is situated on the Turgay trough connecting the West Siberian and Turan plains. In the west, the Turgay trough is bounded by the Trans-Ural Plateau; in the east — by the Kazakh upland and spurs of the Ulutau mountains. Natural and geographical features of Turgay trough allow for their cultural layers to mainly contain diachronous material, with Bestamak serving as an example. Due to this specificity, the collections of stone industry discovered in the monuments of the Turgay trough should be studied on the basis of technical and typological analysis, the main parameters of which being products of primary cleavage; morphological parameters of the plates, the size of plates and tools on the plates; percentage ratio of ingots and tools from plates and rock flakes; secondary processing methods; typological composition of tool kit. The composition of raw materials is used as an independent indicator. In the course of the research, the author concluded that the primary cleavage, nuclei “rejuvenation” and secondary processing of blanks were performed on the site of the settlement. Judging by the results of the technical and typological analysis, the stone industry was deposited from the end of the Mesolithic-the Early Neolithic to the Late Eneolithic. The Neolithic stone industries, starting from the early and late Eneolithic, are predominant at Bestamak. The presence of Mesolithic and Early Eneolithic stone industries on the site is just an assumption. Fragments of ceramics and metal products suggest that people stayed at Bestamak until Late Bronze.

Middle Neolithic of the Kamchatka Peninsula

Purpose. The study of archaeological sites of the Middle Neolithic of Kamchatka should offer a chronology and a set of criteria for identifying the period. Results. The research data is based on the materials of the studied cultural layer, buried dwellings and individual artifacts of 46 archaeological sites. It has been established that the average Neolithic of Kamchatka can be dated back to 4 000–1 500 cal BP. The sites were found on high water-glacial terraces with a height of 4 to 30 meters on the banks of large rivers and lakes, the sea coast of the eastern part of the peninsula. Their number had increased compared to the previous period. Dwellings had become more complex. Perhaps this is due to the need to have more reliable shelters in the conditions of the marine climate and frequent precipitation of volcanic ash. The ground buildings, semi-underground dwellings and workshops for the manufacture of stone tools were found at the sites. Near the dwellings, special fortifications in the form of artificial ditches and ramparts made of stones and soil were also found. These may have been defensive structures. The increased population size, its settlement mainly along the coast in order to develop marine resources, may have caused conflicts between certain groups of the population in the struggle for the best fishing sites. The stone industry is represented by cores (amorphous and prismatic knife-shaped blades) and primary cleavage products (knife-shaped blades of different sizes without retouching, with edge retouching and on both sides). Among the tools there were retouched triangular stone arrowheads without stem and with stem, leaf-shaped, including miniature, arrowheads; knives – narrow and wide-bladed with a dedicated handle, leaf-shaped oval; roughly beaten and polished sharp-edged adzes of different sizes with a sub-triangular and oval cross-section; end scrapers of various geometric shapes; calibrators of arrow shafts. The strategy of life support of society was aimed at hunting for marine mammals, fishing and gathering, including shellfish. In the sphere of spiritual culture, signs of ceremonial activity (labrets) and art (small figurines and ornaments) have also been identified. Conclusion. It is assumed that with an increased population size and changes in the environmental situation, a new way of life of the population developed, associated with a highly specialized and complex appropriating economy which essentially formed its own archaeological culture (Taryinskaya culture).

CUT&RUNTools: a flexible pipeline for CUT&RUN processing and footprint analysis

We introduce CUT&RUNTools as a flexible, general pipeline for facilitating the identification of chromatin-associated protein binding and genomic footprinting analysis from antibody-targeted CUT&RUN primary cleavage data. CUT&RUNTools extracts endonuclease cut site information from sequences of short-read fragments and produces single-locus binding estimates, aggregate motif footprints, and informative visualizations to support the high-resolution mapping capability of CUT&RUN. CUT&RUNTools is available at https://bitbucket.org/qzhudfci/cutruntools/.

CUT&RUNTools: a flexible pipeline for CUT&RUN processing and footprint analysis

We introduce CUT&RUNTools as a flexible, general pipeline for facilitating the identification of chromatin-associated protein binding and genomic footprinting analysis from antibody-targeted CUT&RUN primary cleavage data. CUT&RUNTools extracts endonuclease cut site information from sequences of short read fragments and produces single-locus binding estimates, aggregate motif footprints, and informative visualizations to support the high-resolution mapping capability of CUT&RUN. CUT&RUNTools is available at https://bitbucket.org/qzhudfci/cutruntools/.

Granule proteases of hematopoietic cells, a family of versatile inflammatory mediators – an update on their cleavage specificity, in vivo substrates, and evolution

Cells from several of the hematopoietic cell lineages including mast cells, basophils, neutrophils, cytotoxic T cells, and natural killer (NK) cells store proteases at very high levels within their cytoplasmic granules. In mast cells, these proteases can account for up to 35% of the total cellular protein, and the absolute majority of these belong to the chymotrypsin-related serine protease family. A number of very diverse functions have been identified for these proteases, including apoptosis induction, blood pressure regulation, inactivation of insect and snake toxins, intestinal parasite expulsion, killing of bacteria and fungi, induction, mobilization, or degradation of cytokines, and the degradation of connective tissue components. A very broad spectrum of primary cleavage specificities has also been observed, including chymase, tryptase, asp-ase, elastase, and met-ase specificities, which highlights the large flexibility in the active site of these proteases. Mast cells primarily express chymases and tryptases with chymotryptic or tryptic primary cleavage specificities, respectively. Neutrophils have several enzymes with chymase, elastase, and tryptase specificities. T cells and NK cells express between 5 and 14 different granzymes, depending on the species, and these enzymes have tryptase, asp-ase, chymase, and met-ase specificities. This review focuses on the appearance of these proteases during vertebrate evolution, their primary and extended cleavage specificities, and their potential in vivo substrates. The in vivo substrates and functions are a particular challenging issue because several of these enzymes have a relatively broad specificity and may therefore cleave a wide range of different substrates.

In vivo contribution of serine proteases to the proteolytic activation of γENaC in aldosterone-infused rats

Aldosterone plays an important role in the regulation of blood pressure by modulating the activity of the epithelial sodium channel (ENaC) that consists of α-, β-, and γ-subunits. Aldosterone induces a molecular weight shift of γENaC from 85 to 70 kDa that is necessary for the channel activation. In vitro experiments demonstrated that a dual cleavage mechanism is responsible for this shift. It has been postulated that furin executes the primary cleavage in the Golgi and that the second cleavage is provided by other serine proteases such as prostasin or plasmin at the plasma membrane. However, the in vivo contribution of serine proteases to this cleavage remains unclear. To address this issue, we administered the synthetic serine protease inhibitor camostat mesilate (CM) to aldosterone-infused rats. CM decreased the abundance of the 70-kDa form of ENaC and led to a new 75-kDa form with a concomitant increase in the urinary Na-to-K ratio. Because CM inhibits the protease activity of serine proteases such as prostasin and plasmin, but not furin, our findings strongly indicate that CM inhibited the second cleavage of γENaC and subsequently suppressed ENaC activity. The results of our current studies also suggest the possibility that the synthetic serine protease inhibitor CM might represent a new strategy for the treatment of salt-sensitive hypertension in humans.

Reconstitution of human azurocidin catalytic triad and proteolytic activity by site-directed mutagenesis

Azurocidin belongs to the serprocidin family, but it is devoid of proteolytic activity due to a substitution of His and Ser residues in the catalytic triad. The aim of this study was to reconstitute the active site of azurocidin by site-directed mutagenesis, analyze its processing and restored proteolytic activity. Azurocidin expressed inSf9 insect cells possessing the reconstituted His41-Asp89-Ser175 triad exhibited significant proteolytic activity toward casein with a pH optimum of approximately 8–9, but a reconstitution of only one active site amino acid did not result in proteolytically active protein. Enzymatically active recombinant azurocidin caused cleavage of the C-terminal fusion tag with the primary cleavage site after lysine at Lys-Leu and after alanine at Ala-Ala, and the secondary cleavage site after arginine at Arg-Gln, as well as with low efficiency caused cleavage of insulin chain B after leucine at Leu-Tyr and Leu-Cys, and after alanine at Ala-Leu. We demonstrate that cleavage of the azurocidin C-terminal tripeptide is not necessary for its enzymatic activity. The first isoleucine present in mature azurocidin can be replaced by similar amino acids, such as leucine or valine, but its substitution by histidine or arginine decreases proteolytic activity.

Antiplasmin-cleaving enzyme is a soluble form of fibroblast activation protein

Circulating antiplasmin-cleaving enzyme (APCE) has a role in fibrinolysis and appears structurally similar to fibroblast activation protein (FAP), a cell-surface proteinase that promotes invasiveness of certain epithelial cancers. To explore this potential relationship, we performed comparative structure/function analyses of the 2 enzymes. APCE from human plasma and recombinant FAP (rFAP) exhibited identical pH optima of 7.5, extinction coefficients (\batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \({\in}_{280\mathrm{nm}}^{1\%}\) \end{document}) of 20.2 and 20.5, common sequences of tryptic peptides, and cross-reactivity with FAP antibody. APCE and rFAP are homodimers with monomeric subunits of 97 and 93 kDa. Only homodimers appear to have enzymatic activity, with essentially identical kinetics toward Met-α2-antiplasmin (Met-α2AP) and peptide substrates. APCE and rFAP cleave both Pro3-Leu4 and Pro12-Asn13 bonds of Met-α2AP, but relative kcat/Km values for Pro12-Asn13 are about 16-fold higher than for Pro3-Leu4. APCE and rFAP demonstrate higher kcat/Km values toward a peptide modeled on P4-P4′ sequence surrounding the Pro12-Asn13 primary cleavage site than for Z-Gly-Pro-AMC and Ala-Pro-AFC substrates. These data support APCE as a soluble derivative of FAP and Met-α2AP as its physiologic substrate. Conversion of Met-α2AP by membrane or soluble FAP to the more easily fibrin-incorporable form, Asn-α2AP, may increase plasmin inhibition within fibrin surrounding certain neoplasms and have an impact on growth and therapeutic susceptibility.