QM/MM Description of Newly Selected Catalytic Bioscavengers Against Organophosphorus Compounds Revealed Reactivation Stimulus Mediated by Histidine Residue in the Acyl-Binding Loop

Butyrylcholinesterase (BChE) is considered as an efficient stoichiometric antidote against organophosphorus (OP) poisons. Recently we utilized combination of calculations and ultrahigh-throughput screening (uHTS) to select BChE variants capable of catalytic destruction of OP pesticide paraoxon. The purpose of this study was to elucidate the molecular mechanism underlying enzymatic hydrolysis of paraoxon by BChE variants using hybrid quantum mechanical/molecular mechanical (QM/MM) calculations. Detailed analysis of accomplished QM/MM runs revealed that histidine residues introduced into the acyl-binding loop are always located in close proximity with aspartate residue at position 70. Histidine residue acts as general base thus leading to attacking water molecule activation and subsequent SN2 inline hydrolysis resulting in BChE reactivation. This combination resembles canonical catalytic triad found in active centers of various proteases. Carboxyl group activates histidine residue by altering its pKa, which in turn promotes the activation of water molecule in terms of its nucleophilicity. Observed re-protonation of catalytic serine residue at position 198 from histidine residue at position 438 recovers initial configuration of the enzyme’s active center, facilitating next catalytic cycle. We therefore suggest that utilization of uHTS platform in combination with deciphering of molecular mechanisms by QM/MM calculations may significantly improve our knowledge of enzyme function, propose new strategies for enzyme design and open new horizons in generation of catalytic bioscavengers against OP poisons

Отношение к скорости социальных процессов: разработка новой методики и оценка её валидности

Introduction. The article is devoted to the current but understudied problem in psychology: the speed of social processes. The interdisciplinary approach (theories of P. Virilio, H. Rosa) is used when discussing the issue, substantiating the purpose and hypotheses of the study. The article is aimed to propose the author’s Attitudes towards the Speed of Social Processes inventory, determine its psychometric properties and test its validity. The attitude towards speed is considered as an aspect of subjective time. Methods. The sample size was 521 people. The average age was 31.5 years (min – 21, max – 45), 48.8% of them were men, and 65.6% with higher education; the sample included advanced workers and graduates of vocational educational institutions (work experience 2-3 years). To assess convergent validity, the following were used: Questionnaire of Attitudes towards Technology by G. U. Soldatova, T. A. Nestik, E. I. Rasskazova, E. A. Dorokhova; Personal Flexibility at the Labour Sphere Scale by A. N. Diomin, O. V. Kireeva; scales measuring attitudes towards remote technologies. To assess the criterion validity, the graduates of vocational educational institutions and advancing-age workers were compared (age criterion). Exploratory and confirmatory factor analysis, Spearman’s ρ correlation coefficient and the Mann-Whitney U-test were used. Results. The structure of the inventory is set apart and confirmed. It includes two scales: awareness of the social acceleration (the cognitive component) and rejection of the social acceleration (the affective component); their internal and retest reliability is acceptable. The scales correlate with technophilia, technophobia, technopessimism, attitude to remote technologies, and flexibility of the individual at the labour sphere. It has been established that graduates of vocational educational institutions demonstrate a significantly higher level of awareness and emotional acceptance of social acceleration compared to advanced-age workers. Discussion. The correlations and differences expected in theoretical terms are empirically confirmed. The conclusion is made: the Attitudes Towards the Speed of Social Processes inventory is a new compact psychodiagnostic tool that can be used in psychological and interdisciplinary research. Ideas are formulated that aim to expand the list of criteria for the validity of the new inventory.Введение. Статья посвящена актуальной, но малоизученной в психологии проблеме скорости социальных процессов. Используется междисциплинарный подход (теории П. Вирильо, Х. Розы) при обсуждении проблемы, обосновании цели и гипотез исследования. Цель статьи – предложить авторскую методику «Отношение к скорости социальных процессов», определить её психометрические свойства, проверить валидность. Отношение к скорости рассматривается как аспект субъективного времени. Методы. Объём выборки составил 521 чел., средний возраст 31,5 года (min – 21, max – 45), из них 48,8% мужчин, 65,6% имеют высшее образование; выборка включала зрелых работников и выпускников профессиональных учебных заведений (стаж работы 2–3 года). Для оценки конвергентной валидности применялись: опросник отношения к технологиям Г. У. Солдатовой, Т. А. Нестика, Е. И. Рассказовой, Е. А. Дорохова; шкала гибкости личности в трудовой сфере А. Н. Дёмина, О. В. Киреевой; шкалы, измеряющие отношение к дистанционным технологиям. Для оценки критериальной валидности сравнивались выпускники профессиональных учебных заведений и работники зрелого возраста (возрастной критерий). Использовались эксплораторный и конфирматорный факторный анализ, коэффициент корреляции ρ Спирмена, U-критерий Манна-Уитни. Результаты. Выделена и подтверждена структура опросника. Он включает две шкалы: осознание социального ускорения (когнитивный компонент) и неприятие социального ускорения (аффективный компонент); их внутренняя и ретестовая надёжность приемлемые. Шкалы коррелируют с технофилией, технофобией, технопессимизмом, отношением к дистанционным технологиям, гибкостью личности в трудовой сфере. Установлено, что выпускники профессиональных учебных заведений демонстрируют существенно более высокий уровень осознания и эмоционального приятия социального ускорения по сравнению с работниками зрелого возраста. Обсуждение результатов. Теоретически ожидаемые взаимосвязи и различия эмпирически подтверждены. Сделан вывод: опросник «Отношение к скорости социальных процессов» является новым компактным психодиагностическим инструментом, который можно использовать в психологических и междисциплинарных исследованиях. Формулируются идеи, направленные на расширение перечня критериев валидности новой методики

ИЗУЧЕНИЕ ТВЁРДЫХ ДИСПЕРСИЙ ФУРАЗОЛИДОНА РЕНТГЕНО-ФАЗОВЫМ МЕТОДОМ

Background: to study the phase composition of the solid dispersion of furazolidone by the X-ray phase method. Method: The study was carried out on the basis of the All-Russian Research Institute of Aviation Materials («VIAM») on an X-ray diffractometer DRON-4 («Burevestnik», Russia) according to OFS 1.2.1.1.0011.15 (state Pharmacopoeia XIV). The initial substance of furazolidone, polyvinylpyrrolidone-10000 (PVP) and their solid dispersions (SD) were studied. Result: the radiograph of the SD is the sum of the peaks of the components. SD, presumably, is a combined system - a solution of furazolidone in a polymer and a colloidal phase of the active substance distributed in a matrix of PVP (solid colloid). Conclusion: the obtained data confirm the assumption that furazolidone loses its crystal structure when its SD is obtained with the polymer under study.Цель: изучить фазовый состав твёрдой дисперсии фуразолидона рентгено-фазовым методом. Метод: Исследование проводили на базе ФГУП Всероссийского НИИ авиационных материалов («ВИАМ») на рентгеновском дифрактометре ДРОН-4 (НПП «Буревестник», Россия) согласно ОФС 1.2.1.1.0011.15 (ГФ XIV). Исследовали исходную субстанцию фуразолидона, поливинилпирролидон-10000 (ПВП) и их твёрдые дисперсии (ТД). Результат: рентгенограмма ТД является суммой пиков компонентов. ТД, предположительно, представляет собой комбинированную систему - раствор фуразолидона в полимере и коллоидную фазу действующего вещества, распределённую в матрице ПВП (твёрдый коллоид). Выводы: полученные данные подтверждают предположение о потере фуразолидоном кристаллической структуры при получении его ТД с исследуемым полимером

ИЗУЧЕНИЕ ТВЁРДЫХ ДИСПЕРСИЙ индометацина МЕТОДОМ МИКРОСКОПИИ

Background: to study the effect of obtaining solid dispersions (SD) with polyvinylpyrrolidone-10000 (PVP) on the microcrystalline pattern of indomethacin. Methods: the analysis was carried out at the Department of Analytical, Physical and Colloidal Chemistry of the A.P. Nelyubin Institute of Pharmacy of the I.M. Sechenov First Moscow State Medical University of the Ministry of Health of Russia (Sechenov University). We used a Levenhuk D50L NG digital microscope (made by Levenhuk, China), equipped with a digital camera (2 megapixels) for microphotography, with Levenhuk ToupView software compatible with Windows 7. Substance indomethacin were studied by optical microscopy under a cover glass in a drop of paraffin oil. Under microscopy, recrystallized substance, PVP, etc., a drop of their solution in 96% ethanol was applied to the slide. Microscopy after removal of the solvent. Result: The SD of indomethacin with PVP is a homogeneous system. SD is a solution of indomethacin in a polymer matrix of PVP. Conclusion: the preparation of SD reduces the crystallinity of ndomethaci, improving its dissolution in water.Цель: изучить влияние получения твёрдых дисперсий (ТД) с поливинилпирролидоном-10000 (ПВП) на микрокристаллическую картину индометацина. Метод: анализ проводили на кафедре аналитической, физической и коллоидной химии Института фармации им. А.П. Нелюбина Первый МГМУ им. И.М. Сеченова Минздрава России (Сеченовский Университет). Использовали цифровой микроскоп Levenhuk D50L NG (фирмы Levenhuk, Китай), оснащённый цифровой камерой (2 Мпикс) для микрофотосъёмки, с программным обеспечением Levenhuk ToupView, совместимым с Windows 7. Субстанцию индометацина микроскопировали под покровным стеклом в капле вазелинового масла. При микроскопии, перекристаллизованной субстанции, ПВП и ТД на предметное стекло наносили каплю их раствора в этаноле 96%. Микроскопировали после удаления растворителя. Результат: ТД индометацина с ПВП представляет собой гомогенную систему. ТД - это раствор индометацина в полимерной матрице ПВП. Выводы: получение ТД снижает кристалличность индометацина, улучшая его растворение в воде

ИЗУЧЕНИЕ ОПТИЧЕСКИХ СВОЙСТВ РАСТВОРОВ ТВЁРДОЙ ДИСПЕРСИИ МЕТРОНИДАЗОЛА

Background: to study the effect of obtaining solid dispersions (SD) on the optical properties of metronidazole solutions. Methods: the studied solutions were filtered through Minisart syringe nozzles (Satorius, Germany) with a nylon membrane filter and a pore size of 0.45 microns. The filtered samples were placed in a quartz cuvette (layer thickness 50.0 mm) with the corresponding solution through a hole on the side with a diameter of ≈1 mm (in a light-tight partition between the light source and the wall of the cuvette), a concentrated beam of light was directed. With the help of a Canon 5D MarkII SLR camera, digital images of the Faraday-Tyndall «cone» were taken in a darkened room (exposure time of 20 seconds). Result: opalescence in the form of a bluish-gray cone is observed in solutions of DD metronidazole. Conclusion: the observed Faraday-Tyndall effect confirms the assumptions about the colloidal-dispersed state of metronidazole in the TD solution.Цель: изучить влияние получения твёрдых дисперсий (ТД) на оптические свойства растворов метронидазола. Метод: исследуемые растворы фильтровали через шприцевые насадки Minisart (Satorius, Германия) с мембранным фильтром из нейлона и размером пор 0,45 мкм. Фильтрованные образцы помещали в кварцевую кювету (толщина слоя 50,0 мм) с соответствующим раствором через отверстие сбоку с диаметром ≈1 мм (в светонепроницаемой перегородке между источником света и стенкой кюветы) направляли концентрированный луч света. С помощью зеркальной камеры Canon 5D MarkII в затемнённом помещении делали цифровые изображения «конуса» Фарадея-Тиндаля (время экспозиции 20 сек). Результат: в растворах ТД метронидазола наблюдается опалесценция в виде конуса синевато-серого оттенка. Выводы: наблюдаемый эффект Фарадея-Тиндаля подтверждает предположения о коллоидно-дисперсном состоянии метронидазола в растворе ТД

Intranasal Ion-Triggered In Situ Delivery System of Virus-like Particles: Development Using the Quality by Design Approach

The rapid growth in the prevalence of infectious diseases requires timely action from drug developers. In recent years, the COVID-19 pandemic has demonstrated the unpreparedness of the population for such emergencies. The introduction of modern methods of Design of Experiments (DoE) is required to accelerate the process of drug development and bring a drug to market. The main objective of this study was to develop an ion-triggered in situ system for intranasal delivery of VLP using a Quality by Design approach. Based on a literature review and initial studies, the key QTPP, CQA, CPP, and CMA were identified to develop a novel delivery system for virus-like particles. As a result of the studies on the quality attributes of the developed delivery system, an ion-triggered in situ gel meeting all the specified parameters was obtained using the Quality by Design method

Synthesis of Magneto-Controllable Polymer Nanocarrier Based on Poly(N-isopropylacrylamide-co-acrylic Acid) for Doxorubicin Immobilization

In this work, the preparation procedure and properties of anionic magnetic microgels loaded with antitumor drug doxorubicin are described. The functional microgels were produced via the in situ formation of iron nanoparticles in an aqueous dispersion of polymer microgels based on poly(N-isopropylacrylamide-co-acrylic acid) (PNIPAM-PAA). The composition and morphology of the resulting composite microgels were studied by means of X-ray diffraction, Mössbauer spectroscopy, IR spectroscopy, scanning electron microscopy, atomic-force microscopy, laser microelectrophoresis, and static and dynamic light scattering. The forming nanoparticles were found to be β-FeO(OH). In physiological pH and ionic strength, the obtained composite microgels were shown to possess high colloid stability. The average size of the composites was 200 nm, while the zeta-potential was −27.5 mV. An optical tweezers study has demonstrated the possibility of manipulation with microgel using external magnetic fields. Loading of the composite microgel with doxorubicin did not lead to any change in particle size and colloidal stability. Magnetic-driven interaction of the drug-loaded microgel with model cell membranes was demonstrated by fluorescence microscopy. The described magnetic microgels demonstrate the potential for the controlled delivery of biologically active substances

Role of κ→λ light-chain constant-domain switch in the structure and functionality of A17 reactibody

The engineering of catalytic function in antibodies requires precise information on their structure. Here, results are presented that show how the antibody domain structure affects its functionality. The previously designed organophosphate-metabolizing reactibody A17 has been re-engineered by replacing its constant κ light chain by the λ chain (A17λ), and the X-ray structure of A17λ has been determined at 1.95 Å resolution. It was found that compared with A17κ the active centre of A17λ is displaced, stabilized and made more rigid owing to interdomain interactions involving the CDR loops from the VL and VH domains. These VL/VH domains also have lower mobility, as deduced from the atomic displacement parameters of the crystal structure. The antibody elbow angle is decreased to 126° compared with 138° in A17κ. These structural differences account for the subtle changes in catalytic efficiency and thermodynamic parameters determined with two organophosphate ligands, as well as in the affinity for peptide substrates selected from a combinatorial cyclic peptide library, between the A17κ and A17λ variants. The data presented will be of interest and relevance to researchers dealing with the design of antibodies with tailor-made functions

Presentation_1_QM/MM Description of Newly Selected Catalytic Bioscavengers Against Organophosphorus Compounds Revealed Reactivation Stimulus Mediated by Histidine Residue in the Acyl-Binding Loop.PDF

<p>Butyrylcholinesterase (BChE) is considered as an efficient stoichiometric antidote against organophosphorus (OP) poisons. Recently we utilized combination of calculations and ultrahigh-throughput screening (uHTS) to select BChE variants capable of catalytic destruction of OP pesticide paraoxon. The purpose of this study was to elucidate the molecular mechanism underlying enzymatic hydrolysis of paraoxon by BChE variants using hybrid quantum mechanical/molecular mechanical (QM/MM) calculations. Detailed analysis of accomplished QM/MM runs revealed that histidine residues introduced into the acyl-binding loop are always located in close proximity with aspartate residue at position 70. Histidine residue acts as general base thus leading to attacking water molecule activation and subsequent SN2 inline hydrolysis resulting in BChE reactivation. This combination resembles canonical catalytic triad found in active centers of various proteases. Carboxyl group activates histidine residue by altering its pK_a, which in turn promotes the activation of water molecule in terms of its nucleophilicity. Observed re-protonation of catalytic serine residue at position 198 from histidine residue at position 438 recovers initial configuration of the enzyme’s active center, facilitating next catalytic cycle. We therefore suggest that utilization of uHTS platform in combination with deciphering of molecular mechanisms by QM/MM calculations may significantly improve our knowledge of enzyme function, propose new strategies for enzyme design and open new horizons in generation of catalytic bioscavengers against OP poisons.</p