Dissociation Mechanics and Stability of Type a Botulinum Neurotoxin Complex by Means of Biophysical Evaluation

Biophysical characterization of type A botulinum neurotoxin (BoNT/A) complex along with its thermodynamic stability was assessed through a combination of various methods. BoNT/A exists as large complexes in association with neurotoxin associated proteins (NAPs). To evaluate its biophysical behavior, size-exclusion chromatography (SEC), multi-angled light scattering (MALS), enzyme linked immunosorbent assay (ELISA), and dynamic light scattering (DLS) were utilized. Initially, a single peak (peak 1) of SEC was observed at pH 6.0, and an additional peak (peak 2) appeared at pH 7.4 with a decrement of peak 1. Through MALS and ELISA, the peak 2 was determined to be BoNT/A dissociated from its complex. The dissociation was accelerated by time and temperature. At 37°C, dissociated BoNT/A self-associated at pH 7.4 in the presence of polysorbate 20. On the other hand, the dissociation was partly reversible when titrated back to pH 6.0. Overall, BoNT/A was more stable when associated with NAPs at pH 6.0 compared to its dissociated state at pH 7.4. The conventional analytical methods could be utilized to relatively quantify its amount in different formulations.

pH-Sensitive Glycyrrhizin Based Vesicles for Nifedipine Delivery

Glycyrrhizic acid, or glycyrrhizin (GA), a major active component of licorice root, has been widely used in traditional Chinese and Japanese medicine since ancient times. However, only in the last decades has a novel and unusual property of the GA been discovered to form water-soluble, supramolecular complexes with a variety of lipophilic drugs. These complexes show significant advantages over other known delivery systems, in particular, due to strong pH sensitivity, the properties of GA self-associates. In the present study, a supramolecular complex formation of the hypotensive and antiarrhythmic drug nifedipine with GA has been studied at different pH values, corresponding to the different degrees of GA dissociation, including a fully dissociated state of GA. Both NMR experiments and molecular dynamics simulations demonstrate the existence of the nifedipine complex with GA at all dissociation states of GA. However, optical absorption experiments show the decrease of complex stability and solubility at pH > 6 when the GA molecule is fully deprotonated. It means the higher release rate of the drug in a neutral and basic environment compared with acid media. These results could form the basis of follow-up studies of GA self-associates as pH-controlled drug delivery systems.

Incidence of Emergence Agitation in Pediatric Patient after General Anesthesia

Post anesthesia agitation is common problem in pediatric post anesthesia care unit. The incidences range from 10 to 80%. EA has been described as a dissociated state of consciousness in which the child is inconsolable, irritable, and uncooperative typically thrashing, crying, moaning or incoherent. This study was done to determine the incidence of emergence agitation and associated risk factors in pediatric patients who underwent general anesthesia. This descriptive and analytic study was performed on 105 pediatric patient aged 1-12 years that underwent general anesthesia for various elective diagnostic and surgeries at Dr. Soetomo Hospital between January and February 2016. The presence of emergence agitation was recorded using Pediatric Anesthesia Emergence Delirium (PAED) scale. The factors that linked with Emergence Agitation were recorded in a questionnaire. The data were analyzed using SPSS software with logistic regression. p - values less than 0.05 were considered as significant. Forty two (40%) children had Emergence Agitation. Preoperative anxiety (p = 0.006) and Pain (p=0.035) were associated with higher rates of post anesthetic emergence agitation. This study identified preoperative anxiety and pain as risk factors, which are associated with emergence agitation in children. To minimize the incidence of post anesthetic emergence agitation, these risk factors should be considered in the routine care by anesthetist.

Nightmare Disorder and Isolated Sleep Paralysis

Nightmare disorder and recurrent isolated sleep paralysis are rapid eye movement (REM) parasomnias that cause significant distress to those who suffer from them. Nightmare disorder can cause insomnia due to fear of falling asleep through dread of nightmare occurrence. Hyperarousal and impaired fear extinction are involved in nightmare generation, as well as brain areas involved in emotion regulation. Nightmare disorder is particularly frequent in psychiatric disorders and posttraumatic stress disorder. Nonmedication treatment, in particular imagery rehearsal therapy, is especially effective. Isolated sleep paralysis is experienced at least once by up to 40% of the general population, whereas recurrence is less frequent. Isolated sleep paralysis can be accompanied by very intense and vivid hallucinations. Sleep paralysis represents a dissociated state, with persistence of REM atonia into wakefulness. Variations in circadian rhythm genes might be involved in their pathogenesis. Predisposing factors include sleep deprivation, irregular sleep–wake schedules, and jetlag. The most effective therapy consists of avoiding those factors.

Marked structural rearrangement of mannose 6-phosphate/IGF2 receptor at different pH environments

Many cell surface receptors internalize their ligands and deliver them to endosomes, where the acidic pH causes the ligand to dissociate. The liberated receptor returns to the cell surface in a process called receptor cycling. The structural basis for pH-dependent ligand dissociation is not well understood. In some receptors, the ligand binding domain is composed of multiple repeated sequences. The insulin-like growth factor 2 receptor (IGF2R) contains 15 β strand–rich repeat domains. The overall structure and the mechanism by which IGF2R binds IGF2 and releases it are unknown. We used cryo-EM to determine the structures of the IGF2R at pH 7.4 with IGF2 bound and at pH 4.5 in the ligand-dissociated state. The results reveal different arrangements of the receptor in different pH environments mediated by changes in the interactions between the repeated sequences. These results have implications for our understanding of ligand release from receptors in endocytic compartments.

Dissociation Behavior of Dislocations in Ice

Dislocations in ice behave very differently from those in other materials due to the very low energies of stacking faults in the ice basal plane. As a result, the dislocations dissociate on the basal plane, from a perfect dislocation into two partial dislocations with equilibrium width we ranging from 20 to 500 nm, but what is the timescale to reach this dissociated state? Using physical models, we estimate this timescale by calculating two time-constants: the dissociation-completing time td and the dissociation-beginning time tb. These time constants are calculated for two Burgers vectors as a function of temperature. For perfect dislocations with Burgers vector <c + a>, td is more than one month even at the melting temperature TM, and it exceeds 103 years below −50 ℃, meaning that the dissociation cannot be completed during deformation over laboratory timescales. However, in this case the beginning time tb is less than one second at TM, and it is within several tens of minutes above −50 ℃. These dislocations can glide on non-basal planes until they turn to the dissociated state during deformation, finally resulting in sessile extended dislocations of various widths approaching to the equilibrium value we. In contrast, for perfect dislocations with Burgers vector <a>, td is less than one second above −50 ℃, resulting in glissile extended dislocations with the equilibrium width we on the basal plane. This width is sensitive to the shear stress τ exerted normal to the dislocation line, leading to extension of the intervening stacking fault across the entire crystal grain under commonly accessible stresses. Also, due to the widely dissociated state, dislocations <a> cannot cross-slip to non-basal planes. Such behavior of extended dislocations in ice are notable when compared to those of other materials.

Incidence of Emergence Agitation in Pediatric Patient after General Anesthesia

Post anesthesia agitation is common problem in pediatric post anesthesia care unit. The incidences range from 10 to 80%. EA has been described as a dissociated state of consciousness in which the child is inconsolable, irritable, and uncooperative typically thrashing, crying, moaning or incoherent. This study was done to determine the incidence of emergence agitation and associated risk factors in pediatric patients who underwent general anesthesia. This descriptive and analytic study was performed on 105 pediatric patient aged 1-12 years that underwent general anesthesia for various elective diagnostic and surgeries at Dr. Soetomo Hospital between January and February 2016. The presence of emergence agitation was recorded using Pediatric Anesthesia Emergence Delirium (PAED) scale. The factors that linked with Emergence Agitation were recorded in a questionnaire. The data were analyzed using SPSS software with logistic regression. p - values less than 0.05 were considered as significant. Forty two (40%) children had Emergence Agitation. Preoperative anxiety (p = 0.006) and Pain (p=0.035) were associated with higher rates of post anesthetic emergence agitation. This study identified preoperative anxiety and pain as risk factors, which are associated with emergence agitation in children. To minimize the incidence of post anesthetic emergence agitation, these risk factors should be considered in the routine care by anesthetist.

An intermediate state of bacterial ribosome dissociation is fixed by yeast mitochondrial initiation factor 3

The processes of association and dissociation of ribosomal subunits are of great importance for the protein biosynthesis. The mechanistic details of these processes, however, are not well known. In bacteria, upon translation termination, ribosome dissociates into subunits which is necessary for its further involvement into new initiation step. The dissociated state of ribosome is maintained by initiation factor 3 (IF3) which binds to free small subunits and prevents their premature association with the large subunits. In this work, we have exchanged IF3 in E.coli cells by its ortholog from Saccharomyces cerevisiae mitochondria (Aim23p) and showed that yeast protein cannot functionally substitute the bacterial one and is even slightly toxic for bacterial cells. Our in vitro experiments have demonstrated that Aim23p does not split E.coli ribosomes into subunits. Instead, it fixes an intermediate state of ribosomes dissociation characterized by sedimentation coefficient about 60S. Using molecular modeling, we show that such fixation is due to mitochondria-specific terminal extensions of Aim23p that stabilize the position of the protein on the bacterial ribosome.

An intermediate state of bacterial ribosome dissociation is fixed by yeast mitochondrial initiation factor 3

The processes of association and dissociation of ribosomal subunits are of great importance for the protein biosynthesis. The mechanistic details of these processes, however, are not well known. In bacteria, upon translation termination, ribosome dissociates into subunits which is necessary for its further involvement into new initiation step. The dissociated state of ribosome is maintained by initiation factor 3 (IF3) which binds to free small subunits and prevents their premature association with the large subunits. In this work, we have exchanged IF3 in E.coli cells by its ortholog from Saccharomyces cerevisiae mitochondria (Aim23p) and showed that yeast protein cannot functionally substitute the bacterial one and is even slightly toxic for bacterial cells. Our in vitro experiments have demonstrated that Aim23p does not split E.coli ribosomes into subunits. Instead, it fixes an intermediate state of ribosomes dissociation characterized by sedimentation coefficient about 60S. Using molecular modeling, we show that such fixation is due to mitochondria-specific terminal extensions of Aim23p that stabilize the position of the protein on the bacterial ribosome.

Estimation of Polyelectrolyte Phase Volume Based on the Analysis of Acid Dissociation Equilibia of Polyacrylic Acid

The acid dissociation equilibria of typical linear weak acid polyelectrolytes, polyacrylic acid (PAA), were investigated by a potentiometric titration method. The electrostatic effects on the equilibria have been rationalized based on a Gibbs-Donnan concept. The apparent acid dissociation constant of PAA at completely neutralized condition determined by extrapolation are quite close to acid dissociation constant of their monomeric acid. The effective volume and the radius of polyelectrolyte phase of PAA have been calculated experimentally by the non-ideality term of ΔpK values with an iterative procedure. The radiuses increase with increasing at α 0.2 region. It is attributable to the repulsion between negatively charged carboxylate groups is weak at lowly dissociated state, whereas the repulsion is strong in highly dissociated state.