Secreted proteins of eukaryotes are decorated with branched carbohydrate oligomers called glycans. This fact is only starting to be considered for
investigations of protein dynamics. Using all-atom ...molecular dynamics (MD) simulations and Markov state modeling (MSM), we unveil the influence of glycans on the conformational flexibility of the multidomain protein disulfide isomerase (PDI), which is a ubiquitous chaperone in the endoplasmic reticulum (ER). Yeast PDI (yPDI) from
is glycosylated at asparagine side chains and the knowledge of its five modified sites enables a realistic computational modeling. We compare simulations of glycosylated and unglycosylated yPDI and find that the presence of glycan-glycan and glycan-protein interactions influences the flexibility of PDI in different ways. For example, glycosylation reduces interdomain interactions, shifting the conformational ensemble toward more open, extended structures. In addition, we compare our results on yPDI with structural information of homologous proteins such as human PDI (hPDI), which is natively unglycosylated. Interestingly, hPDI lacks a surface recess that is present in yPDI. We find that glycosylation of yPDI facilitates its catalytic site to reach close to this surface recess. Hence, this might point to a possible functional relevance of glycosylation in yeast to act on substrates, while glycosylation seems redundant for the human homologous protein. We conclude that glycosylation is fundamental for protein dynamics, making it a necessity for a truthful representation of the flexibility and function in
studies of glycoproteins.
Ligand–receptor binding and unbinding are fundamental biomolecular processes and particularly essential to drug efficacy. Environmental water fluctuations, however, impact the corresponding ...thermodynamics and kinetics and thereby challenge theoretical descriptions. Here, we devise a holistic, implicit-solvent, multi-method approach to predict the (un)binding kinetics for a generic ligand–pocket model. We use the variational implicit-solvent model (VISM) to calculate the solute–solvent interfacial structures and the corresponding free energies, and combine the VISM with the string method to obtain the minimum energy paths and transition states between the various metastable (“dry” and “wet”) hydration states. The resulting dry–wet transition rates are then used in a spatially dependent multistate continuous-time Markov chain Brownian dynamics simulation and the related Fokker–Planck equation calculations of the ligand stochastic motion, providing the mean first-passage times for binding and unbinding. We find the hydration transitions to significantly slow down the binding process, in semiquantitative agreement with existing explicit-water simulations, but significantly accelerate the unbinding process. Moreover, our methods allow the characterization of nonequilibrium hydration states of pocket and ligand during the ligand movement, for which we find substantial memory and hysteresis effects for binding vs. unbinding. Our study thus provides a significant step forward toward efficient, physics-based interpretation and predictions of the complex kinetics in realistic ligand–receptor systems.
Along with the growing geriatric population, the number of odontoid fractures is steadily increasing. However, the effectiveness of immobilizing geriatric odontoid fractures using a cervical collar ...has been questioned. The aim of the present study is to analyze the physiological and pathological motion in odontoid fractures and to assess limitation of motion in the cervical spine when applying a cervical collar.
Motion analysis was performed with wireless motion tracker on unfixed geriatric human cadavers. First, a new geriatric type II odontoid fracture model was developed. In this model, the type II odontoid fracture is operated via a transoral approach. The physiological and pathological flexion and lateral bending of the cervical spine resulting from this procedure was measured. The resulting motion after external stabilization using a cervical collar was analyzed.
The new geriatric type II odontoid fracture model was successfully established using seven unfixed human cadavers. The pathological flexion of the cervical spine was significantly increased compared to the physiological flexion (p = 0.027). Furthermore, the flexion was significantly reduced when a cervical collar was applied. In case of flexion the mean remaining motion was significantly reduced (p = 0.0017) from 41° to 14°. For lateral bending the mean remaining motion was significantly reduced (p = 0.0137) from 48° to 18°.
In case of type II odontoid fracture, flexion and lateral bending of the cervical spine are increased due to spinal instability. Thus, if an odontoid fracture is suspected in geriatric patients, the application of a cervical collar should always be considered since external stabilization can significantly reduce flexion and lateral bending.
The calculation of relative free-energy differences between different compounds plays an important role in drug design to identify potent binders for a given protein target. Most rigorous methods ...based on molecular dynamics simulations estimate the free-energy difference between pairs of ligands. Thus, the comparison of multiple ligands requires the construction of a “state graph”, in which the compounds are connected by alchemical transformations. The computational cost can be optimized by reducing the state graph to a minimal set of transformations. However, this may require individual adaptation of the sampling strategy if a transformation process does not converge in a given simulation time. In contrast, path-free methods like replica-exchange enveloping distribution sampling (RE-EDS) allow the sampling of multiple states within a single simulation without the pre-definition of alchemical transition paths. To optimize sampling and convergence, a set of RE-EDS parameters needs to be estimated in a pre-processing step. Here, we present an automated procedure for this step that determines all required parameters, improving the robustness and ease of use of the methodology. To illustrate the performance, the relative binding free energies are calculated for a series of checkpoint kinase 1 inhibitors containing challenging transformations in ring size, opening/closing, and extension, which reflect changes observed in scaffold hopping. The simulation of such transformations with RE-EDS can be conducted with conventional force fields and, in particular, without soft bond-stretching terms.
The aim of this study was to compare the remaining motion of an immobilized cervical spine using an innovative cervical collar as well as two traditional cervical collars. The study was performed on ...eight fresh human cadavers. The cervical spine was immobilized with one innovative (Lubo Airway Collar) and two traditional cervical collars (Stifneck and Perfit ACE). The flexion and lateral bending of the cervical spine were measured using a wireless motion tracker (Xsens). With the Weinman Lubo Airway Collar attached, the mean remaining flexion was 20.0 ± 9.0°. The mean remaining flexion was lowest with the Laerdal Stifneck (13.1 ± 6.6°) or Ambu Perfit ACE (10.8 ± 5.8°) applied. Compared to that of the innovative Weinmann Lubo Airway Collar, the remaining cervical spine flexion was significantly decreased with the Ambu Perfit ACE. There was no significant difference in lateral bending between the three examined collars. The most effective immobilization of the cervical spine was achieved when traditional cervical collars were implemented. However, all tested cervical collars showed remaining motion of the cervical spine. Thus, alternative immobilization techniques should be considered.
We investigate how to tune the rate of hydrophobic ligand-receptor association due to the role of solvent in adjustable receptor pockets by explicit-water molecular dynamics (MD) simulations. Our ...model considers the binding of a spherical ligand (key/guest) to a concave surface recess in a nonpolar wall as receptor (lock/host). We systematically modify the receptor's physicochemical properties in terms of geometry and dispersion attraction which, in turn, alter the water occupancy and fluctuations within the pocket. We demonstrate that even minor pocket modifications can lead to a significant acceleration of the water-mediated association. For example, the binding switches from comparably slow to fast if the binding pocket becomes only slightly deeper. We find that the degree of hydrophobicity, characterized by hydration occupancy and its fluctuations, clearly correlates with the binding times and, for instance, links the sudden acceleration to an abrupt increase in hydrophobicity. For a deeper analysis based on passage time theory, we quantify the intimate coupling between solvent fluctuations and the ligand's local dynamics and friction. The coupling exhibits substantial nonequilibrium effects and maximizes shortly before binding, which slows down the binding kinetics in all cases. In summary, we rationalize how the physicochemical properties of a nonpolar, concave binding site tune key-lock binding kinetics due to water-mediated forces and fluctuations. Our study thus complements the profound understanding of the solvent's influence in host-guest binding, which is essential for tailored solutions in catalysis and pharmaceutical applications.
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