Penicillin acylase catalyzed synthesis of penicillin-G from substrates anchored in cyclodextrins

Penicillin acylase (EC 3.5.1.11) catalyses the condensation of phenylacetic acid (PAA) and 6-aminopenicillanic acid (6-APA) to form benzylpenicillin (BP). Both PAA and 6-APA were found to form host-guest complexes with beta-methylcyclodextrin (beta m-CD) and gamma-cyclodextrin (gamma-CD) respectively. The rate of the reaction catalyzed by the enzyme remained unaffected if one of the substrates used was in the cyclodextrin complexed form. However, in this case, the reaction lasted longer and yielded about 20 per cent more products compared to the condensation reaction involving only uncomplexed substrates. There was distinct increase in the rate of formation of the antibiotic, if both substrates used are in CD-complexed form.     

Comparison of two trunk electromagnetic sensor placement methods during shoulder motion analysis

For kinematic studies of the shoulder, electromagnetic sensors are commonly placed on the humerus, scapula, and trunk. The trunk sensor is used to describe humeral and scapular kinematics with respect to the trunk. There are two common trunk sensor placements, the sternum or third thoracic vertebrae (T3). It is currently unclear if placement of the trunk sensor affects kinematics, making it difficult to compare data across studies. The purpose of this study was to compare two trunk sensor placements (T3 and sternum) on trunk and scapular kinematics during arm elevation. An electromagnetic tracking system was used to collect kinematic data during five consecutive repetitions of ascending and descending arm elevation in the sagittal plane. The results indicate that trunk sensor placement had no significant effect on trunk kinematics or scapular upward/downward rotation and internal/external rotation. Scapular anterior/posterior tilt was significantly greater when the trunk sensor was on the sternum compared to the T3 vertebrae during ascending 30°–120°: mean difference = −3.51° (95%CI: −5.61, −1.40), and descending 120°–30°: mean difference = −3.27° (95%CI: −6.07, −0.48). However, the difference in anterior/posterior tilt did not exceed the error (minimal detectable change), and thus is likely not a meaningful difference. These results indicate the trunk sensors can be affixed on T3 or the sternum, depending on the needs of the study.     

Evaluation of a flexible force sensor for measurement of helmet foam impact performance

The association between translational head acceleration and concussion remains unclear and provides a weak predictive measure for this type of injury; thus, alternative methods of helmet evaluation are warranted. Recent finite element analysis studies suggest that better estimates of concussion risk can be obtained when regional parameters of the cranium, brain and surrounding tissues are included. Lacking, however, are empirical data at the head–helmet interface with regards to contact area and force. Hence, the purpose of this study was to evaluate a system to capture the impact force distribution of helmet foams. Thirteen Flexiforce^® sensors were arranged in a 5×5 cm array, secured to a load cell. Three densities of foam were repeatedly impacted with 5 J of energy during ambient (20 °C) and cold (?25 °C) conditions. RMS error, calculated relative to the global force registered by the load cell, was <1.5% of the measurement range during individual calibration of the Flexiforce^® sensors. RMS error was 5% of the measured range for the global force estimated by the sensor array. Load distribution measurement revealed significant differences between repeated impacts of cold temperature foams for which acceleration results were non-significant. The sensor array, covering only 36% of the total area, possessed sufficient spatial and temporal resolution to capture dynamic load distribution patterns. Implementation of this force mapping system is not limited to helmet testing. Indeed it may be adopted to assess other body regions vulnerable to contact injuries (e.g., chest, hip and shin protectors).     

A model for the shuttle motions of puerarin and daidzin inside the cavity of β-cyclodextrin in aqueous acetic acid: insights from molecular dynamics simulations

Acetic acid acts as one component of the mobile phase to influence separation of puerarin from daidzin when using β-cyclodextrin-substituted media. In this work considering an explicit acetic acid solution, host-guest complexes of β-cyclodextrin (β-CD) with puerarin and daidzin were investigated by molecular dynamics simulations. Computational results indicate different shuttle motions of puerarin and daidzin inside the cavity of β-CD. A model detailing the shuttle motion was constructed, and the relationships between shuttle depth and guest rotation angles, hydrogen bonds, and host-guest interaction energies were analyzed. The results can be used to explain the chromatographic retention mechanisms of puerarin and daidzin with β-CD, and to explore the complexity of host-guest interactions involving β-CD.     

A DNA duplex-based, tailor-made fluorescent sensor for porphyrin derivatives

We synthesized a DNA-based fluorescent sensor, tailored to the recognition and signaling for hydrophobic guest molecules. The sensor molecule is made of three functional moieties, per-O-methylated beta-cyclodextrin (per-Me-beta-CD), DNA, and pyrene as guest-binding, dimerizing, and signaling ones, respectively. Addition of a porphyrin derivative as a hydrophobic guest molecule induced emission switching of the sensor molecule from monomer to excimer due to the DNA hybridization promoted by the host-guest binding.     

A structural model of polyglutamine determined from a host-guest method combining experiments and landscape theory

Modeling the structure of natively disordered peptides has proved difficult due to the lack of structural information on these peptides. In this work, we use a novel application of the host-guest method, combining folding theory with experiments, to model the structure of natively disordered polyglutamine peptides. Initially, a minimalist molecular model (C(alpha)C(beta)) of CI2 is developed with a structurally based potential and captures many of the folding properties of CI2 determined from experiments. Next, polyglutamine "guest" inserts of increasing length are introduced into the CI2 "host" model and the polyglutamine is modeled to match the resultant change in CI2 thermodynamic stability between simulations and experiments. The polyglutamine model that best mimics the experimental changes in CI2 thermodynamic stability has 1), a beta-strand dihedral preference and 2), an attractive energy between polyglutamine atoms 0.75-times the attractive energy between the CI2 host Go-contacts. When free-energy differences in the CI2 host-guest system are correctly modeled at varying lengths of polyglutamine guest inserts, the kinetic folding rates and structural perturbation of these CI2 insert mutants are also correctly captured in simulations without any additional parameter adjustment. In agreement with experiments, the residues showing structural perturbation are located in the immediate vicinity of the loop insert. The simulated polyglutamine loop insert predominantly adopts extended random coil conformations, a structural model consistent with low resolution experimental methods. The agreement between simulation and experimental CI2 folding rates, CI2 structural perturbation, and polyglutamine insert structure show that this host-guest method can select a physically realistic model for inserted polyglutamine. If other amyloid peptides can be inserted into stable protein hosts and the stabilities of these host-guest mutants determined, this novel host-guest method may prove useful to determine structural preferences of these intractable but biologically relevant protein fragments.     

Measuring Enthalpy of Sublimation of Volatiles by Means of Piezoelectric Crystal Microbalances

Piezoelectric Crystal Microbalances (PCM’s) are widely used to study the chemical processes involving volatile compounds in any environment, such as condensation process. Since PCM’s are miniaturized sensor, they are very suitable for planetary in situ missions, where can be used to detect and to measure the mass amount of astrobiologically significant compounds, such as water and organics. This work focuses on the realization and testing of a new experimental setup, able to characterize volatiles which can be found in a planetary environment. In particular the enthalpy of sublimation of some dicarboxylic acids has been measured. The importance of dicarboxylic acids in planetology and astrobiology is due to the fact that they have been detected in carbonaceous chondritic material (e.g. Murchinson), among the most pristine material present in our Solar System. In this work, a sample of acid was heated in an effusion cell up to its sublimation. For a set of temperatures (from 30 °C to 75 °C), the deposition rate on the PCM surface has been measured. From these measurements, it has been possible to infer the enthalpy of sublimation of Adipic acid, i.e. ΔH = 141.6 ± 0.8 kJ/mol and Succinic acid, i.e. ΔH = 113.3 ± 1.3 kJ/mol. This technique has so demonstrated to be a good choice to recognise a single compound or a mixture (with an analysis upstream) even if some improvements concerning the thermal stabilization of the system will be implemented in order to enhance the results’ accuracy. The experiment has been performed in support of the VISTA (Volatile In Situ Thermogravimetry Analyzer) project, which is included in the scientific payload of the ESA MarcoPolo-R mission study.     

Paramagnetic NMR Investigation of Dendrimer-Based Host-Guest Interactions

In this study, the host-guest behavior of poly(amidoamine) (PAMAM) dendrimers bearing amine, hydroxyl, or carboxylate surface functionalities were investigated by paramagnetic NMR studies. 2,2,6,6-Tetramethylpiperidinyloxy (TEMPO) derivatives were used as paramagnetic guest molecules. The results showed that TEMPO-COOH significantly broaden the ¹H NMR peaks of amine- and hydroxyl-terminated PAMAM dendrimers. In comparison, no paramagnetic relaxation enhancement (PRE) was observed between TEMPO-NH₂, TEMPO-OH and the three types of PAMAM dendrimers. The PRE phenomenon observed is correlated with the encapsulation of TEMPO-COOH within dendrimer pockets. Protonation of the tertiary amine groups within PAMAM dendrimers plays an important role during this process. Interestingly, the absence of TEMPO-COOH encapsulation within carboxylate-terminated PAMAM dendrimer is observed due to the repulsion of TEMPO-COO- anion and anionic dendrimer surface. The combination of paramagnetic probes and ¹H NMR linewidth analysis can be used as a powerful tool in the analysis of dendrimer-based host-guest systems.     

Temperature and relative humidity effects on egg and nymphal development of Aceria tulipae (K.) (Acari: Eriophyidae) on garlic leaves (Allium sativum L.)

Aceria tulipae (K.) is responsible for important crop losses of garlic in all production areas around the world. However, very little is known about its development in relation to temperature and relative humidity (r.h). Laboratory rearings from egg to adult stage were done on pieces of the apical part of young garlic leaves. These leaf pieces were placed in aluminium dishes floating on water or put on a solid hygroscopic salt in closed plastic trays. Rearing done in dark incubators at constant temperatures (10°C to 45°C) and with different r.h (25% to 75%) at 15°C and 25°C showed that: i) the shortest development time occurs at about 25°C; ii) the upper lethal temperature and the development threshold for the eggs are respectively estimated about 45°C and 6°C; iii) a r.h close to 100% is required for a high percentage of egg hatching, but water condensation on leaves is harmful. Host transpiration has an important regulatory effect on humidity conditions favouring mite survival.     

On-line evaluation of fermentation broth colour

Summary A novel method for evaluating pigment production during fermentation ofMonascus ruber has been developed and consists of reflectance measurement with a 10° observer under an artificial illuminant. A spectrophotometer was adapted to measure pigmentation during culture in a bioreactor. The lightness parameter L was the most useful information delivered by this sensor. An absorbency over 30 units at 480nm on batch cultures was estimated on-line without dilution. member of the GR Automatique - SARTA - CNRS     

Platinum Layers Sandwiched Between Black Phosphorous and Graphene for Enhanced SPR Sensor Performance

Highly sensitive surface plasmon resonance (SPR) sensor consisting of Ag-Pt bimetallic films sandwiched with 2D materials black phosphorus (BP) and graphene over Pt layer in Kretschmann configuration is analyzed theoretically using the transfer matrix method. Numerical results show that upon suitable optimization of thickness of Ag-Pt layers and the number of layers of BP and graphene, sensitivity as high as 412°/RIU (degree/refractive index unit) can be achieved for p-polarized light of wavelength 633 nm. This performance can be tuned and controlled by changing the number of layers of BP and graphene. Furthermore, the addition of graphene and heterostructures of black phosphorus not only improved the sensitivity of the sensor but also kept the FWHM of the resonance curve much smaller than the conventional sensor utilizing Au as plasmonic metal and hence improved the resolution to a significant extent. We expect that this new proposed design will be useful for medical diagnosis, biomolecular detection, and chemical examination.

     

Local secondary structure in ribonuclease A denatured by guanidine · HCl near 1 °C

Recent work has shown that a short α-helix can be stable in water near 1 °C when stabilized by specific interactions between side-chains, while earlier “host-guest” results with random copolymers have shown that a short α-helix is unstable in water at all temperatures in the absence of stabilizing side-chain interactions. As regards the mechanism of protein folding, it is now reasonable on energetic grounds to consider isolated α-helices and β-sheets as the first intermediates on the pathway of protein folding. Proton nuclear magnetic resonance is used here to detect isolated secondary structures in ribonuclease A denatured by guanidine · HCl (GuHCl). Temperatures near 1 °C are used because the low-temperature stability of the C-peptide helix may be a general property of isolated secondary structures in water.Our procedure is to titrate with GuHCl the C2H resonance lines of the four histidine residues of denatured ribonuclease A. Studies of model peptides (C-peptide (lactone) and C-peptide carboxylate, residues 1 to 13 of ribonuclease A; S-peptide, residues 1 to 20) show linear titration curves for the C2H resonance of His12 above 0.5 M-GuHCl, once helix unfolding is complete. Deviations from this line are used to monitor helix formation. The GuHCl titration curves of the other three histidine residues are also linear, once unfolding is complete. The results show that the helix found in C-peptide and S-peptide is also found in denatured ribonuclease A, where it behaves as an isolated helix not stabilized significantly by interactions with other chain segments. Studies of denatured S-protein show that the remaining three His residues, His48, His105 and His119, are involved in structure only below 1 m-GuHCl at 9 °C, pH 1.9. The nature of this structure is not known. The main conclusion from this work is that the His12 helix can be observed as a stable, isolated helix in denatured ribonuclease A near 1 °C, and that none of the other three His residues is involved in a comparably stable local structure. In native ribonuclease A, His12 is within an α-helix and the other three His residues are involved in a 3-stranded β-sheet structure.The helix-coil transition of C-peptide has also been studied for other side-chain resonances by GuHCl titration. Typically, but not always, the titration curves are linear after helix unfolding takes place and resonance lines from different residues of the same amino acid type can be resolved in GuHCl solutions. This is true of the four histidine residues of ribonuclease A although their pK values in 5 m-GuHCl are nearly the same. In C-peptide, the βCH₃ resonance of Ala6 is affected strongly by GuHCl while the lines of Ala4 and Ala5 are shifted only weakly by GuHCl. Evidently the interactions between GuHCl and side-chains in an unfolded peptide depend upon neighboring groups.     

Temperature and developmental responses of body and cell size in Drosophila; effects of polyploidy and genome configuration

Increased adult body size in Drosophila raised at lower temperatures could be attributed both to an increase in the cell volume and cell number. It is not clear, however, whether increased cell size is related to (or even caused by) increased nuclear volume and genome size (or configuration). Experiments with Drosophila melanogaster stocks (Oregon-R and w1118) raised at 16, 22, 24, and 28 °C resulted in larger adult body and wing size with lower temperature, while eye size was less affected. The increase in wing size reflected an increase in cell size in both males and females of both stocks. The nucleus size, genome size, and DNA condensation of adult flies, embryos, and Schneider 2 cells (S2 cells, of larval origin) were estimated by flow cytometry. In both adult flies and S2 cells, both nucleus size and DNA condensation varied with temperature, while DNA content appears to be constant. From 12% to 18% of the somatic cells were tetraploid (4C) and 2–5% were octoploid (8C), and for the Oregon strain we observed an increase in the fraction of polyploid cells with decreasing temperature. The observed increase in body size (and wing size) at low temperatures could partly be linked with the cell size and DNA condensation, while corresponding changes in the haploid genome size were not observed.     

A topical metabolic inhibitor to improve transcutaneous estimation of arterial oxygen tension in adults

The use of Eugenol as a contact medium in the measurement of transcutaneous oxygen (tcPO₂) partially inhibits the metabolism in the underlying tissue, thereby reducing oxygen consumption and increasing tcPO₂. Oxygen consumption in the tissue can be estimated from the rate at which tcPO₂ falls when blood flow is occluded, and blood flow in the tissue can be estimated from the rate at which tcPO₂ increases when the subject changes from breathing air to pure oxygen. Both these measurements have been made with Eugenol and distilled water as contact media. From these measurements it has proved possible to estimate the arterial oxygen tension (aPO₂) of healthy adults at a relatively low sensor temperature (43°C).     

Validity and reliability of a shoe-embedded sensor module for measuring foot progression angle during over-ground walking

Wearable systems are becoming increasingly popular for gait assessment outside of laboratory settings. A single shoe-embedded sensor module can measure the foot progression angle (FPA) during walking. The FPA has important clinical utility, particularly in populations with knee osteoarthritis, as it is a target for biomechanical treatments. However, the validity and the day-to-day reliability of FPA measurement using wearable systems during over-ground walking has yet to be established. Two gait analysis sessions on 20 healthy adults were conducted. During both sessions, participants performed natural over-ground walking in a motion capture laboratory and on a 100 m linear section of outdoor athletics track. FPA was measured in the laboratory via marker trajectory data, while the sensor module measured FPA during the outdoor track walking. Validity was examined by comparing the laboratory- and sensor-measured average FPA. Day-to-day reliability was examined by comparing the sensor-measured FPA between the first and second gait analysis sessions. Average absolute error between motion capture and sensor measured FPA were 1.7° and 2.1° at session 1 and 2, respectively. A Bland and Altman plot indicated no systematic bias, with 95% limit of agreement widths of 4.2° – 5.1°. Intraclass correlation coefficient (ICC_2k) analysis resulted in good to excellent validity (ICC = 0.89 – 0.91) and reliability (ICC = 0.95). Overall, the shoe-embedded sensor module is a valid and reliable method of measuring FPA during over-ground walking without the need for laboratory equipment.     

The Condensation of DNA by Chromium (III) Ions

Abstract

Cr(III), one of the most potent inorganic carcinogens, induces condensation of DNA into a very compact product at 37°, as shown by electron microscopy. The condensation begins with the appearing of some supercoil structures and complete condensation occurs at relatively low Cr(III) concentrations; for 3 and 30 mM ionic strength they are 4.5 and 45 μM, respectively. Under these conditions, Cr(III) inhibits the interaction between ethidium and DNA as shown by absorption and fluorescence spectra.     

Thermal porosity analysis of croscarmellose sodium and sodium starch glycolate by differential scanning calorimetry

The aim of the study was to demonstrate the applicability of differential scanning calorimetry (DSC) on porosity analysis for cellulose and starch. Croscarmellose sodium (CCS) and sodium starch glycolate (SSG) were allowed to sorb moisture in 85%, 90%, 95%, and 100% relative humidity (RH) at 40°C for 24 hours. The pretreated samples were then subjected to DSC running temperature ranging from 25°C to −50°C at a cooling rate of 10°C/min. The cooling traces of water crystallization, if present, were transformed to porosity distribution via capillary condensation using Kelvin's equation. The porosity analysis of CCS and SSG was also done using nitrogen adsorption as a reference method. It was found that sorbed water could not be frozen (in cases of 85% and 90% RH) until the moisture content exceeded a cutoff value (in cases of 95% and 100% RH). The nonfreezable moisture content was referred to tightly bound, plasticizing water, whereas the frozen one may be attributed to loosely bound water condensation in pore structure of CCS and SSG surfaces. Not only capillary condensation but also the tightly bound, nonfreezable monolayer water lying along the inner pores of the surface contributed to porosity determination. Good agreement with less than 5% deviation of mean pore size was observed when the results were compared with nitrogen adsorption. The narrower pore size distributions, however, were obtained because of the limitations of the technique. It was concluded that pore analysis by DSC could be successful. Further research needs to be done to account for limitations and to extend the applicability of the technique.     

Thermal sensitivity of a Neotropical amphibian (Engystomops pustulosus) and its vulnerability to climate change

A species’ thermal sensitivity and its exposure to climate variation are key components in the prediction of its vulnerability to climate change. We tested the thermal sensitivity of a tropical amphibian that lives in a mild constant climate in which the thermal tolerance range is expected to closely match the experienced environmental temperature. The air temperature that this species is exposed to varies between 21.9 and 31.6°C with an annual mean of 27.2°C. We estimated the microhabitat water temperature variation under vegetation shade, which buffers the temperature by 1.8°C in relation to that of the air, and with open canopy, where the water was 1.9°C warmer than the air temperature. With broods of tadpoles split into five treatments (15°C, 21°C, 28°C, 31°C, and 33°C), we estimated the critical thermal maximum (CTMax) and critical thermal minimum (CTMin) after at least 7 days of acclimation. Both CTMax (42.3°C) and CTMin (11.8°C) were more extreme than the temperature range estimated for the field. We estimated the optimum temperature (T_o = 28.8°C) and the thermal performance breadth (range: 23.3–34.1°C) based on growth rate (g/day). The animals were able to acclimate more extensively to cold than to warm temperatures. These performance curve traits closely matched the air temperature. The estimated vulnerability varied according to the microhabitat prediction model used. The combination of tadpole data on thermal sensitivity and macro‐ and microhabitat variation provides a necessary framework to understand the effects of climate change on tropical amphibians.