62 research outputs found

    Determining The Site Specific Metal Binding and Structural Properties of EF-Hand Protein Using Grafting Approach

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    Calmodulin is an essential EF-hand protein with a helix-loop-helix calcium binding motif. Understanding Ca(II) dependent activation of calmodulin and other EF-hand proteins is limited by Ca(II)-induced conformational change, multiple and cooperative binding of Ca(II) ions, and interactions between the paired EF-hand motifs. The goal of this research project is to probe key determinants for calcium binding properties and pairing interactions at the site specific level using a grafting approach and high resolution NMR. An individual Ca(II) binding site of the EF-hand motifs of calmodulin was grafted into a non-calcium dependent protein, CD2, to bypass limitations associated with natural EF-hand proteins and peptide fragments. Using high resolution NMR, we have shown that the grafted EF-loop III of calmodulin in the host protein retains its native conformation with a strong loop and β-conformation preference. Grafted ligand residues in the engineered protein are directly involved in binding of Ca(II) and La(III). The NMR studies support our hypothesis that both ligand arrangement and dynamic properties play essential role in tuning Ca(II) binding affinities. Using pulse-field diffusion NMR and protein engineering, we further demonstrated that grafted EF- loop remains as a monomer. Although the EF-loop with flanking helices dimerizes in the presence of Ca(II). Additionally, removal of conserved hydrophobic residues at the flanking helices of the EF-hand motif leads to be monomer in the absence and presence of metal ions. Our results suggest that conserved hydrophobic residues are essential for the pair-paired interaction in the coupled EF-hand protein. We have shown that our developed grafting approach can be applied to probe intrinsic Ca(II) binding affinities of different Ca(II) binding sites

    Theoretical Study of Chloroperoxidase Catalyzed Chlorination of beta-Cyclopentanedione and Role of Water in the Chlorination Mechanism

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    Chloroperoxidase (CPO) is a potential biocatalyst for use in asymmetric synthesis. The mechanisms of CPO catalysis are therefore of interest. The halogenation reaction, one of several chemical reactions that CPO catalyzes, is not fully understood and is the subject of this dissertation. The mechanism by which CPO catalyzes halogenation is disputed. It has been postulated that halogenation of substrates occurs at the active site. Alternatively, it has been proposed that hypochlorous acid, produced at the active site via oxidation of chloride, is released prior to reaction, so that halogenation occurs in solution. The free-solution mechanism is supported by the observation that halogenation of most substrates often occurs non-stereospecifically. On the other hand, the enzyme-bound mechanism is supported by the observation that some large substrates undergo halogenation stereospecifically. The major purpose of this research is to compare chlorination of the substrate beta-cyclopentanedione in the two environments. One study was of the reaction with limited hydration because such a level of hydration is typical of the active site. For this work, a purely quantum mechanical approach was used. To model the aqueous environment, the limited hydration environment approach is not appropriate. Instead, reaction precursor conformations were obtained from a solvated molecular dynamics simulation, and reaction of potentially reactive molecular encounters was modeled with a hybrid quantum mechanical/molecular mechanical approach. Extensive work developing parameters for small molecules was pre-requisite for the molecular dynamics simulation. It is observed that a limited and optimized (active-site-like) hydration environment leads to a lower energetic barrier than the fully solvated model representative of the aqueous environment at room temperature, suggesting that the stable water network near the active site is likely to facilitate the chlorination mechanism. The influence of the solvent environment on the reaction barrier is critical. It is observed that stabilization of the catalytic water by other solvent molecules lowers the barrier for keto-enol tautomerization. Placement of water molecules is more important than the number of water molecules in such studies. The fully-solvated model demonstrates that reaction proceeds when the instantaneous dynamical water environment is close to optimal for stabilizing the transition state

    Prevalence of GBV-C RNA in HIV infected individuals in Tehran, Iran

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    Background: GB virus C (GBV-C) is a flavivirus that was characterized in 1995. The prevalence of GBV-C RNA in HIVinfected subjects has not been previously studied in Iran and was therefore determined. Methods: We have investigated serum samples of 80 patients from Imam Khomeini Hospital, Tehran, Iran, whose HIV infection was confirmed in our laboratory by Western-blotting. We used nested-PCR to detect GBV-C/HGV RNA in their sera. Results: GBV-C/HGV RNA was detected in 15 (18.8) of 80 patients. There was no significant difference in GBV-C/HGV RNA prevalence between males and females, in different age groups and HIV viral loads groups. Conclusion: The prevalence of GBV-C RNA was high in HIV-infected patients. There was no association between GBV-C RNA prevalence and specific gender, age, and HIV viral loads groups

    Σφαιρική οπτική ανίχνευση συμβάντων με χρήση του αλγορίθμου APIT

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    Η παρούσα εργασία επικεντρώνεται στον προσδιορισμό της γεωγραφικής θέσης συμβάντων που εντοπίζονται σε εικονορροές. Προτείνεται μια παραλλαγή του αλγορίθμου APIT, που χρησιμοποιείται για χωροθέτηση κόμβων σε ασύρματα δίκτυα αισθητήρων, και παρουσιάζεται ένα σύστημα αυτόματου εντοπισμού που χρησιμοποιεί τον αλγόριθμο αυτό. Ο αλγόριθμος με βάση τις συντεταγμένες του σημείου της εικόνας που προσπαθεί να προσδιορίσει χωρικά και μιας βάσης δεδομένων με σημεία αναφοράς δίνει μια εκτίμηση της γεωγραφικής θέσης του συμβάντος. Επειδή η διαδικασία αυτή είναι ανεξάρτητη για κάθε εικόνα έγιναν πολλές δοκιμές πάνω σε στατικές εικόνες ώστε να διαπιστωθεί η εγκυρότητα του αλγορίθμου και να αναλυθεί η ακρίβειά του. Το σύστημα που αναπτύχθηκε έχει σαν κύριο στόχο να παρουσιάσει τις δυνατότητες χρήσης του αλγορίθμου σε εφαρμογές εντοπισμού θέσης συμβάντων σε βίντεο. Ο αλγόριθμος είναι γρήγορος και κατά περιπτώσεις μπορεί να δώσει πολύ ακριβή αποτελέσματα. Η ακρίβειά του επηρεάζεται κυρίως από την μορφολογία του εδάφους και την πυκνότητα των σημείων αναφοράς και αναλόγως την περίπτωση μπορεί να χρειαστεί κάποια ιδιαίτερη παραμετροποίηση για να μπορεί να δώσει όσο το δυνατόν καλύτερα αποτελέσματα.This work focuses on identifying the geographical location of events found in video- streams. We propose a vairiation of the APIT algorithm, used for node localization in wireless sensor networks , and we present an auto-detection system that uses this algorithm. The algorithm based on the image coordinates of the point we are trying to locate and reference points from a database, gives an estimation of the location of the event. Because this process is independent for each image, many tests were performed on static images to determine the validity of the algorithm and to analyze its accuracy. The main purpose of the developed system is to present the possibilities of using the algorithm in applications that require event localization from video streams. The algorithm is fast and on occasion can give very accurate results. Its accuracy is mainly affected by the terrain morphology and the density of the reference points and, depending on the case, some particular parameterization may be needed to give the best possible results

    Пристрій зарядки акумуляторної батареї з розширеним алгоритмом роботи

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    Пояснювальна записка містить: 46 сторінок, 21 рисунок, 18 літературних джерел. Графічна частина роботи містить: блок-схему алгоритму роботи пристрою, структурну, функціональну і принципову електричну схеми. Пояснювальна записка містить три розділи. Перший розділ містить огляд технічної літератури за вибраним напрямком проектування. Другий розділ присвячений розробці алгоритму функціонування і структурної схеми. В третьому розділі, розроблені функціональна і принципова схеми пристрою і виконаний розрахунок основних вузлів принципової схеми. По результатам розробки зроблені висновки

    Avaliação da toxicidade do mercúrio com um sensor potenciométrico

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    Mestrado em Engenharia QuímicaO objetivo deste trabalho consistiu na aplicação de um elétrodo seletivo ao mercúrio, para a deteção do mercúrio na água do mar em níveis de atividade correspondentes à toxicidade aguda na biota. Biodisponibilidade e, consequentemente, a toxicidade do mercúrio no ambiente aquático dependem de sua forma química. O mercúrio inorgânico, é associado à concentração ou atividade dos iões Hg2+. Dois métodos de avaliação da toxicidade do mercúrio em meio salino foram empregados: potenciometria, com os elétrodos de membrana de vidro calcogeneto seletivos para o mercúrio, e um teste biológico baseado na luminescência da bactéria marinha Vibrio fischeri. Os elétrodos seletivos ao mercúrio foram calibrados em soluções com diferentes conteúdos de cloreto e outros ligantes, e em diferentes valores de pH. O sensor respondeu na gama de atividades de mercúrio de 10-4 a 10-23 mol L-1 e com um limite de deteção na água do mar artificial de 2,65×10-24 mol L1. A toxicidade da água do mar artificial enriquecida com mercúrio, foi medida usando bactéria luminescente V. fischeri. Toxicidade aguda do mercúrio nestas condições, corresponde à atividade de 3,72×10-24 mol L-1 (t = 10 min). O sensor potenciométrico com membrana de vidro calcogeneto, pode ser aplicado para a deteção da atividade de mercúrio correspondente à toxicidade aguda, apesar de esta atividade ser muito próxima do limite de detecção do sensor. As duas amostras recolhidas na Ria de Aveiro não apresentaram toxicidade para V. fischeri, assim, foi-lhes adicionado mercúrio. O comportamento de ambos os sensores e da bactéria nas amostras de água do mar real, foi diferente em comparação com as soluções de água do mar artificial, salientando que esta última não é adequada para a calibração. Verificou-se que o limite de deteção do sensor potenciométrico na água real não permitiu a deteção da atividade do mercúrio, correspondente à toxicidade aguda.The objective of this work was application of the mercury-selective electrode to the detection of the mercury in the seawater at the activity levels corresponding to the acute toxicity to the biota. Bioavailability and consequently toxicity of the mercury in the aquatic environment depends on its chemical form. For inorganic mercury it is associated with concentration of Hg2+ ion or activity. Two methods of assessment of mercury toxicity in saline media were employed: potentiometry with the mercury-selective electrode with chalcogenide glass membrane and biologic test based on the luminescence of the marine bacterium Vibrio fischeri. Mercury-selective electrodes were calibrated in the solutions with different content of chloride and other mercury complexing ligands and different pHs. Sensor has shown response in the range of the mercury activities 10-4 - 10-23 mol L-1 and detection limit in the artificial seawater of 2,65×10-24 mol L-1.Toxicity of artificial seawater spiked with mercury was measured using luminescent bacterium V. fischeri. Acute toxicity of mercury in this conditions corresponds to its activity of 3,72×10-24 mol L-1 (t = 10 min). Potentiometric sensor with chalcogenide glass membrane can be applied to the detection of mercury activity at the level of acute toxicity though this activity is very close to the detection limit of the sensor. Two samples collected in the Ria de Aveiro were found not to be toxic to V. Fischeri, thus they were spiked with mercury. Behavior of both sensor and bacterium in the real seawater samples was different compared to the artificial seawater solutions, stressing that the latter are not adequate for calibration. It was found that detection limit of potentiometric sensor in the real water does not allow detection of mercury on the activity level corresponding to the acute toxicity

    Response of aerospace structures to reentry aerodynamic excitation

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    A bibliography on aerodynamic excitation and associated structural response is presented. Very little applicable data under actual reentry conditions was found, but many wind tunnel tests and some in-flight data were. Aerodynamic excitation and proposed space shuttle configurations and reentry trajectories were studied. Attached turbulent boundary layer, separated turbulent boundary layer, and base pressure fluctuations are the region of aerodynamic excitation most applicable to exterior panels of space shuttle-type vehicles; prediction techniques are presented for these environments. Methods for predicting the response of panel structures to acoustic excitation were investigated. A normal mode approach was developed for the low frequency response and a statistical energy approach was utilized for the high frequency response. A finite element model was made of a thermal protective system test panel. Computer programs were used to determine the natural frequencies and mode shapes of the panel and to calculate the response of the panel to a reverberant acoustic field. Power spectral densities of response displacement and acceleration are presented

    Conformally invariant complete metrics

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    Important geometric or analytic properties of domains in the Euclidean space Rn\mathbb{R}^n or its one-point compactification (the M\"obius space) Rn\overline{\mathbb{R}}^n (n2)(n\ge 2) are often characterized by comparison inequalities between two intrinsic metrics on a domain. For instance, a proper subdomain GG of Rn\mathbb{R}^n is {\it uniform} if and only if the quasihyperbolic metric kGk_G is bounded by a constant multiple of the distance-ratio metric jG.j_G. Motivated by this idea we first characterize the completeness of the modulus metric of a proper subdomain GG of Rn\overline{\mathbb{R}}^n in terms of Martio's MM-condition. Next, we prove that if the boundary is uniformly perfect, then the modulus metric is minorized by a constant multiple of a M\"obius invariant metric which yields a new characterization of uniform perfectness of the boundary of a domain. Further, in the planar case, we obtain a new characterization of uniform domains. In contrast to the above cases, where the boundary has no isolated points, we study planar domains whose complements are finite sets and establish new upper bounds for the hyperbolic distance between two points in terms of a logarithmic M\"obius metric. We apply our results to prove H\"older continuity with respect to the Ferrand metric for quasiregular mappings of a domain in the M\"obius space into a domain with uniformly perfect boundary.Comment: 28 page