All-Atom Molecular Dynamics Investigations on the Interactions between D2 Subunit Dopamine Receptors and Three 11C-Labeled Radiopharmaceutical Ligands.

The D2 subunit dopamine receptor represents a key factor in modulating dopamine release. Moreover, the investigated radiopharmaceutical ligands used in positron emission tomography imaging techniques are known to bind D2 receptors, allowing for dopaminergic pathways quantification in the living human brain. Thus, the biophysical characterization of these radioligands is expected to provide additional insights into the interaction mechanisms between the vehicle molecules and their targets. Using molecular dynamics simulations and QM calculations, the present study aimed to investigate the potential positions in which the D2 dopamine receptor would most likely interact with the three distinctive synthetic 11C-labeled compounds (raclopride (3,5-dichloro-N-[[(2S)-1-ethylpyrrolidin-2-yl]methyl]-2-hydroxy-6-methoxybenzamide)-RACL, FLB457 (5-bromo-N-[[(2S)-1-ethylpyrrolidin-2-yl]methyl]-2,3-dimethoxybenzamide)-FLB457 and SCH23390 (R(+)-7-Chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine)-SCH)), as well as to estimate the binding affinities of the ligand-receptor complexes. A docking study was performed prior to multiple 50 ns molecular dynamics productions for the ligands situated at the top and bottom interacting pockets of the receptor. The most prominent motions for the RACL ligand were described by the high fluctuations of the peripheral aliphatic -CH3 groups and by its C-Cl aromatic ring groups. In good agreement with the experimental data, the D2 dopamine receptor-RACL complex showed the highest interacting patterns for ligands docked at the receptor's top position.

Analytical Perspectives in the Study of Polyvalent Interactions of Free and Surface-Bound Oligonucleotides and Their Implications in Affinity Biosensing.

The high affinity and/or selectivity of oligonucleotide-mediated binding offers a myriad of therapeutical and analytical applications, whose rational design implies an accurate knowledge of the involved molecular mechanisms, concurring equilibrium processes and key affinity parameters. Oligonucleotide-functionalized gold surfaces or nanostructures are regularly employed analytical platforms for the development of label-free optical or electrochemical biosensors, and recently, novel detection platform designs have been increasingly considering the synergistic effect of polyvalent binding, involving the simultaneous interaction of two or several oligonucleotide strands. Considering the general lack of studies involving ternary single-stranded DNA (ssDNA) interactions, a complementary analytical workflow involving capillary gel electrophoretic (CGE) mobility shift assay, microcalorimetry and computational modeling has been deployed for the characterization of a series of free and surface-bound binary and ternary oligonucleotide interactions. As a proof of concept, the DNA analogue of MicroRNA 21 (miR21), a well-known oncogenic short MicroRNA (miRNA) sequence, has been chosen as a target molecule, simulating limiting-case scenarios involved in dual molecular recognition models exploited in affinity (bio)sensing. Novel data for the characterization of oligonucleotide interacting modules is revealed, offering a fast and complete mapping of the specific or non-specific, often competing, binary and ternary order interactions in dynamic equilibria, occurring between various free and metal surface-bound oligonucleotides.

Decreased Interactions between Calmodulin and a Mutant Huntingtin Model Might Reduce the Cytotoxic Level of Intracellular Ca2+: A Molecular Dynamics Study.

Mutant huntingtin (m-HTT) proteins and calmodulin (CaM) co-localize in the cerebral cortex with significant effects on the intracellular calcium levels by altering the specific calcium-mediated signals. Furthermore, the mutant huntingtin proteins show great affinity for CaM that can lead to a further stabilization of the mutant huntingtin aggregates. In this context, the present study focuses on describing the interactions between CaM and two huntingtin mutants from a biophysical point of view, by using classical Molecular Dynamics techniques. The huntingtin models consist of a wild-type structure, one mutant with 45 glutamine residues and the second mutant with nine additional key-point mutations from glutamine residues into proline residues (9P(EM) model). Our docking scores and binding free energy calculations show higher binding affinities of all HTT models for the C-lobe end of the CaM protein. In terms of dynamic evolution, the 9P(EM) model triggered great structural changes into the CaM protein's structure and shows the highest fluctuation rates due to its structural transitions at the helical level from α-helices to turns and random coils. Moreover, our proposed 9P(EM) model suggests much lower interaction energies when compared to the 45Qs-HTT mutant model, this finding being in good agreement with the 9P(EM)'s antagonistic effect hypothesis on highly toxic protein-protein interactions.

Specific Key-Point Mutations along the Helical Conformation of Huntingtin-Exon 1 Protein Might Have an Antagonistic Effect on the Toxic Helical Content's Formation.

The polyglutamine tract length represents a key regulator for the Huntington's disease toxicity level and its aggregation rates, often being related to helical structural conformations. In this study, we performed all-atom MD simulations on mutant Huntingtin-Exon1 protein with additional mutation spots, aiming to observe the corresponding structural and dynamical changes at the level of the helix. The simulated structures consist of three sets of Q residue mutations into P residues (4P, 7P, and 9P), with each set including different spots of mutations: random along the mutant sequence (R models), at the edges of the helix (E models), as well as at the edges and in the middle of the helix (EM models). At the helical level, our results predict less compactness profiles for a higher number of P mutations (7P and 9P models) with particular mutation spots at the edges and at the edges-middle of the helix. Moreover, the C-alpha atom distances decreased for 7P and 9P models in comparison to 4P models, and the RMSF values show the highest fluctuation rates for 9P models with point mutations at the edges and in the middle of the helix. The secondary structure analysis suggests greater structural transitions from α-helices to bends, turns, and random coils for 7P and 9P models, particularly for point mutations considered at the edges and in the middle of the helical content. The obtained results support our hypothesis that specific key-point mutations along the helical conformation might have an antagonistic effect on the toxic helical content's formation.

Molecular Dynamics Simulations Applied to Structural and Dynamical Transitions of the Huntingtin Protein: A Review.

Over the recent years, Huntington's disease (HD) has become widely discussed in the scientific literature especially because at the mutant level there are several contradictions regarding the aggregation mechanism. The specific role of the physiological huntingtin protein remains unknown, due to the lack of characterization of its entire crystallographic structure, making the experimental and theoretical research even harder when taking into consideration its involvement in multiple biological functions and its high affinity for different interacting partners. Different types of models, containing fewer (not more than 35 Qs) polyglutamine residues for the WT structure and above 35 Qs for the mutants, were subjected to classical or advanced MD simulations to establish the proteins' structural stability by evaluating their conformational changes. Outside the polyQ tract, there are two other regions of interest (the N17 domain and the polyP rich domain) considered to be essential for the aggregation kinetics at the mutant level. The polymerization process is considered to be dependent on the polyQ length. As the polyQ tract's dimension increases, the structures present more ß-sheet conformations. Contrarily, it is also considered that the aggregation stability is not necessarily dependent on the number of Qs, while the initial stage of the aggregation seed might play the decisive role. A general assumption regarding the polyP domain is that it might preserve the polyQ structures soluble by acting as an antagonist for ß-sheet formation.