Role of the ectodomain serine 275 in shaping the binding pocket of the ATP-gated P2X3 receptor

ATP-activated P2X3 receptors expressed in nociceptive sensory neurons play an important role in pain signaling. Basic properties of this receptor subtype, including very strong desensitization, depend on the rate of dissociation of the agonist from the binding site. Even though the rough structure of the ATP binding site has been proposed on the basis of the X-ray structure of the zebrafish P2X4 receptor and mutagenesis studies, the fine subunit-specific structural properties predisposing the receptor to tight capture of the agonist inside the binding pocket have not been elucidated. In this work, by exploring in silico the functional role for the left flipper located in the ectodomain region, we identified within this loop a candidate residue S275, which could contribute to the closure of the agonist-binding pocket. Testing of the S275 mutants using the patch-clamp technique revealed a crucial role for S275 in agonist binding and receptor desensitization. The S275A mutant showed a reduced rate of onset of desensitization and accelerated resensitization and was weakly inhibited by nanomolar agonist. Extracellular calcium application produced inhibition instead of facilitation of membrane currents. Moreover, some full agonists became only partial agonists when applied to the S275A receptor. These effects were stronger with the more hydrophobic mutants S275C and S275V. Taken together, our data suggest that S275 contributes to the closure of the agonist-binding pocket and that effective capture of the agonist provided by the left flipper in calcium-dependent manner determines the high rate of desensitization, slow recovery, and sensitivity to nanomolar agonist of the P2X3 receptor. © 2011 American Chemical Society

Highly conserved tyrosine 37 stabilizes desensitized states and restricts calcium permeability of ATP-gated P2X3 receptor

Tyrosine 37 in the first transmembrane (TM1) domain is highly conserved in ATP-gated P2X receptors suggesting its fundamental role. We tested whether Y37 contributes to the desensitization of P2X3 receptors, which is currently not well understood. By combining electrophysiological, imaging and modeling approaches, we studied desensitization of various Y37 P2X3 mutants and potential partners of Y37. Unlike the membrane current of the WT receptor, which desensitized in seconds, Y37A mutant current did not fully desensitize even after minutes-long applications of β,γ-meATP, α,β-meATP, ATP or 2MeS-ATP. The fractional calcium current was enhanced in the Y37A mutant. Y37F did not rescue the native P2X3 phenotype indicating a role for the hydroxyl group of Y37 for the WT receptor. Homology modeling indicated I318 or I319 in TM2 as potential partners for Y37 in the receptor closed state. We tested this hypothesis by creating a permanent interaction between the two residues via disulfide bond. Whereas single Y37C, I318C and I319C mutants were functional, the double mutants Y37C-I318C and Y37C-I319C were non-functional. Using a cyclic model of receptor operation, we suggest that the conserved tyrosine 37 links TM1 to TM2 of adjacent subunit to stabilize desensitized states and restricts calcium permeability through the ion channel. © 2011 International Society for Neurochemistry

Reconstructed Serine 288 in the Left Flipper Region of the Rat P2X7 Receptor Stabilizes Nonsensitized States

© 2017 American Chemical Society. Serine 275, a conserved residue of the left flipper region of ATP-gated P2X3 receptors, plays a key role in both agonist binding and receptor desensitization. It is conserved in most of the P2X receptors except P2X7 and P2X6. By combining experimental patch-clamp and modeling approaches, we explored the role of the corresponding residue in the rat P2X7 receptor (rP2X7) by replacing the phenylalanine at position 288 with serine and characterizing the membrane currents generated by either the wild-type (WT) or the mutated rP2X7 receptor. F288S, an rP2X7 mutation, slowed the deactivation subsequent to 2 and 20 s applications of 1 mM ATP. F288S also prevented sensitization (a progressive current growth) observed with the WT in response to a 20 s application of 1 mM ATP. Increasing the ATP concentration to 5 mM promoted sensitization also in the mutated rP2X7 receptor, accelerating the deactivation rate to typical WT values. YO-PRO1 uptake in cells expressing either the WT or the F288S P2X7 receptor was consistent with recorded membrane current data. Interestingly, in the human P2X7 (hP2X7) receptor, substitution Y288S did not change the deactivation rate, while the Y288F mutant generated a "rat-like" phenotype with a fast deactivation rate. Our combined experimental, kinetic, and molecular modeling data suggest that the rat F288S novel phenotype is due to a slower rate of ATP binding and/or unbinding and stabilization of nonsensitized receptor states

Comparison of open femoral exposure and percutaneous access in endovascular reconstruction of the thoracic aorta: a two-center retrospective study

Aim. To analyze the efficacy and safety of the percutaneous transfemoral puncture technique for TEVAR (thoracis endovascular aortic repair).Material and methods. The retrospective study included 89 patients with aortic pathologies, for whom endovascular repair was performed: 51 patients (57%) with aortic dissection (type I DeBakey — 30 cases (58,8%) and type III — 21 (41,2%)), 38 (43%) patients with aortic aneurism. 82% of patients were male, the median age was 57 years (minimum age 17 years, maximum age 75 years). All patients were divided into two groups: in the first group (48 patients) endovascular aortic repair was performed under endotracheal anesthesia with open femoral exposure of the common femoral artery (CFA), in the second group (41 patients) — by percutaneous puncture method under local anesthesia. Technical and clinical aspects of procedures were analyzed.Results. Technical success of endovascular repair was achieved in 100% cases in both groups. The duration of the operation in the group with percutaneous access was statically significantly shorter (120 (94-150) minutes vs 87(60-120) minutes, p=0,001). Also, the time spent by patients in the intensive care unit and the period of hospitalization (18 (14-22) hours versus 1 (0-3) hours, p=0,001; 5 (4-6) days versus 4 (3-5) days, p=0,03) was shorter. In the open access group 2 (4,2%) patients developed access-related complications - acute thrombosis of the common femoral artery and hematoma of the postoperative wound, which required additional surgical aid - thrombectomy from the CFA, the second patient had evacuation of the hematoma of the postoperative wound. Cite-related complications in the second group were not observed. No major complications including neurological deficits and hospital mortality were observed in both groups.Conclusions. Thoracic endovascular aortic repair (TEVAR) using percutaneous access under local anesthesia in stable patients has proven to be safe and effective. The operation time is significantly reduced and this approach in most cases eliminates the need for the patient to stay in the intensive care unit in the early postoperative period. Possibility of early mobilization of the patient appears with reducing of the duration of hospitalization

Comparative analysis of Illumina and Ion Torrent high-throughput sequencing platforms for identification of plant components in herbal teas

© 2018 Elsevier Ltd The rapid development of high-throughput sequencing (HTS) methods offers new opportunities for food quality control and identification of food components using the DNA barcoding approach (metabarcoding in cases of complex mixes). However, the protocols of DNA barcoding applied to food analysis are not yet fully established; testing and optimization are required to achieve the highest accuracy and cost efficiency. We report here a comparative study of the two most widely used sequencing platforms - Illumina and Ion Torrent - for composition analysis of herbal teas, and show that both technologies yield congruent results, both qualitatively and quantitatively. They have revealed the substitution of fireweed (Epilobium angustifolium L.) by Lythrum sp. in one of the samples. It was confirmed by classic methods of botanical analysis (anatomy and palynology). In most samples, undeclared components have been detected, such as bindweed (Convolvulus) and ragweed (Ambrosia), which are known toxic and allergy-causing plants

Клиническое наблюдение пациента с синхронными опухолями левой почки и правого мочеточника. Описание клинического случая

The paper describes a rare case of synchronous bilateral tumor involvement of the urinary tract.Приводится описание редкого наблюдения больного с синхронным двусторонним опухолевым поражением мочевыделительной системы

A role for diatom-like silicon transporters in calcifying coccolithophores

Biomineralization by marine phytoplankton, such as the silicifying diatoms and calcifying coccolithophores, plays an important role in carbon and nutrient cycling in the oceans. Silicification and calcification are distinct cellular processes with no known common mechanisms. It is thought that coccolithophores are able to outcompete diatoms in Si-depleted waters, which can contribute to the formation of coccolithophore blooms. Here we show that an expanded family of diatom-like silicon transporters (SITs) are present in both silicifying and calcifying haptophyte phytoplankton, including some globally important coccolithophores. Si is required for calcification in these coccolithophores, indicating that Si uptake contributes to the very different forms of biomineralization in diatoms and coccolithophores. Significantly, SITs and the requirement for Si are absent from highly abundant bloom-forming coccolithophores, such as Emiliania huxleyi. These very different requirements for Si in coccolithophores are likely to have major influence on their competitive interactions with diatoms and other siliceous phytoplankton

СПЕКТР МУТАЦИЙ ГЕНА BRCA1 У БОЛЬНЫХ РАКОМ МОЛОЧНОЙ ЖЕЛЕЗЫ В МОЛОДОМ ВОЗРАСТЕ В РОССИИ

Aim of the study. Aim of the study was to estimate the occurrence of pathogenic mutations in the BRCA1 gene in Russian breast cancer patients.Material and methods. Complete coding sequence of the BRCA1 gene of 445 early onset  breast cancer patients (under 40 years) from Novosibirsk region (Russia) were analyzed by  targeted Next Generation Sequencing (NGS) using Ion Torrent platform. Results. Forty (9%) carriers of various pathogenic mutations were revealed. Thirty five (7,9%) patients  carried 5382insC mutation, described earlier as a founder mutation for Slavic population.  Five (1.1%) patients carried various pathogenic mutations, namely C61G, 462delCC, E143X,  4153delA, and IVS18+1G>T. Besides, 29 genetic variants with no clinical significance or with  unknown clinical significance were detected in BRCA1 gene among 445 early onset breast  cancer patients. Conclusions. Data on the frequency of genetic variations in the BRCA1 gene among early onset breast cancer patients in the Novosibirsk Region (Russia) were  obtained. Proportion of the 5382insC mutation is 87.5% of all pathogenic mutations in the BRCA1 gene found in patients.Цель исследования – оценить частоту встречаемости патогенных мутаций в BRCA1 гене у женщин с раком молочной железы, проживающих в России.Материал и методы. Проведён анализ полной кодирующей части гена BRCA1 у 445 больных раком молочной  железы на ранней стадии (возраст больных до 40 лет), проживающих в Новосибирской области (Россия), с  помощью метода таргетного секвенирования на платформе Ion Torrent. Результаты. Выявлено 40 (9 %)  носительниц различных патогенных мутаций. У 35 (7,9 %) пациенток обнаружена мутация 5382insC, описанная  ранее как «мутация-основателя» в славянской популяции. У 5 (1,1 %) пациенток были выявлены  другие различные патогенные мутации, а именно C61G, 462delCC, E143X, 4153delA и IVS18 + 1G> T. Кроме  того, 29 генетических вариантов с отсутствующей или неясной клинической значимостью были обнаружены в  гене BRCA1 у 445 больных раком молочной железы на ранней стадии. Выводы. Получены данные о частоте  генетических вариаций гена BRCA1 у больных раком молочной железы на ранней стадии, проживающих в  Новосибирской области (Россия). Доля мутации 5382insC составляет 87,5 % от всех патогенных мутаций в гене BRCA1, обнаруженных у пациенток

Observation of the radiative decay mode of the free neutron

The theory of quantum electrodynamics (QED) predicts that beta decay of the neutron into a proton, electron and antineutrino should be accompanied by a continuous spectrum of soft photons. While this inner bremsstrahlung branch has been previously measured in nuclear beta and electron capture decay, it has never been observed in free neutron decay. Recently, the photon energy spectrum and branching ratio for neutron radiative decay have been calculated using two approaches: a standard QED framework(1-3) and heavy baryon chiral perturbation theory(4) (an effective theory of hadrons based on the symmetries of quantum chromodynamics). The QED calculation treats the nucleons as point-like, whereas the latter approach includes the effect of nucleon structure in a systematic way. Here we observe the radiative decay mode of free neutrons, measuring photons in coincidence with both the emitted electron and proton. We determined a branching ratio of (3.13 +/- 0.34) x 10(-3) (68 per cent level of confidence) in the energy region between 15 and 340 keV, where the uncertainty is dominated by systematic effects. The value is consistent with the predictions of both theoretical approaches; the characteristic energy spectrum of the radiated photons, which differs from the uncorrelated background spectrum, is also consistent with the calculated spectrum. This result may provide opportunities for more detailed investigations of the weak interaction processes involved in neutron beta decay.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/62639/1/nature05390.pd

Possible neurological complications after carotid endarterectomy: causes and prevention methods

This is a joint research performed by neurologists and vascular surgeons. The aim of this study was to evaluate and compare our outcomes after carotid endarterectomy (CEA) at the vascular surgery department of the University Hospital in Ufa, Russia with the results in other hospitals in the world.Данная работа проведена совместно неврологами и сосудистыми хирургами. Цель настоящего исследования оценить и сравнить результаты каротидной эндартерэктомии (КЗАЗ) в сосудистом отделении клиники Башкирского государственного медицинского университета с мировыми показателям