Dániel Szöllősi

Discrete Molecular Dynamics Can Predict Helical Prestructured Motifs in Disordered Proteins

Intrinsically disordered proteins (IDPs) lack a stable tertiary structure, but their short binding regions termed Pre-Structured Motifs (PreSMo) can form transient secondary structure elements in solution. Although disordered proteins are crucial in many biological processes and designing strategies to modulate their function is highly important, both experimental and computational tools to describe their conformational ensembles and the initial steps of folding are sparse. Here we report that discrete molecular dynamics (DMD) simulations combined with replica exchange (RX) method efficiently samples the conformational space and detects regions populating α-helical conformational states in disordered protein regions. While the available computational methods predict secondary structural propensities in IDPs based on the observation of protein-protein interactions, our ab initio method rests on physical principles of protein folding and dynamics. We show that RX-DMD predicts α-PreSMos with high confidence confirmed by comparison to experimental NMR data. Moreover, the method also can dissect α-PreSMos in close vicinity to each other and indicate helix stability. Importantly, simulations with disordered regions forming helices in X-ray structures of complexes indicate that a preformed helix is frequently the binding element itself, while in other cases it may have a role in initiating the binding process. Our results indicate that RX-DMD provides a breakthrough in the structural and dynamical characterization of disordered proteins by generating the structural ensembles of IDPs even when experimental data are not available.

Comparison of six multiclass classifiers by the use of different classification performance indicators

Classification problems are very important, and generally, the question is which is the best model. Several classification performance indicators including the classification accuracy value (ACC), Cohens kappa (KAPPA), or the area under the ROC curve (AUC) are used to answer this question. There are non‐parametric comparative methods such as the sum of ranking differences method. The objective of this work was to find the best classification method to classify four soft drink samples and four model samples, which differ from each other only in the sweetener composition. Model samples were used to be basic samples for comparison with the commercial soft drinks. Six different classification methods were compared according to their classification performance. A corrected classification accuracy value (corrected ACC) was developed for the purpose and was introduced. This value takes into account the similarities between the classes. The results showed that the ACC value and the KAPPA values give similar results in our case. The best three models according to the ACC, KAPPA, and AUC were “K‐nearest neighbor,” “random forest,” and “discriminant analysis.” However, the corrected ACC value showed a bit different ranking, and the random forest model was neglected from the good models. The confusion matrices of the models confirmed the ranking according to the corrected ACC value. The results showed that the best classification model was the K‐nearest neighbor for the available samples, and the corrected ACC value is a useful classification performance indicator. Copyright

Functional rescue of a misfolded Drosophila melanogaster dopamine transporter mutant associated with a sleepless phenotype by pharmacological chaperones

Folding-defective mutants of the human dopamine transporter (DAT) cause a syndrome of infantile dystonia/parkinsonism. Here, we provide a proof-of-principle that the folding deficit is amenable to correction in vivo by two means, the cognate DAT ligand noribogaine and the HSP70 inhibitor, pifithrin-μ. We examined the Drosophila melanogaster (d) mutant dDAT-G108Q, which leads to a sleepless phenotype in flies harboring this mutation. Molecular dynamics simulations suggested an unstable structure of dDAT-G108Q consistent with a folding defect. This conjecture was verified; heterologously expressed dDAT-G108Q and the human (h) equivalent hDAT-G140Q were retained in the endoplasmic reticulum in a complex with endogenous folding sensors (calnexin and HSP70-1A). Incubation of the cells with noribogaine (a DAT ligand selective for the inward-facing state) and/or pifithrin-μ (an HSP70 inhibitor) restored folding of, and hence dopamine transport by, dDAT-G108Q and hDAT-G140Q. The mutated versions of DAT were confined to the cell bodies of the dopaminergic neurons in the fly brain and failed to reach the axonal compartments. Axonal delivery was restored, and sleep time was increased to normal length (from 300 to 1000 min/day) if the dDAT-G108Q-expressing flies were treated with noribogaine and/or pifithrin-μ. Rescuing misfolded versions of DAT by pharmacochaperoning is of therapeutic interest; it may provide opportunities to remedy disorders arising from folding-defective mutants of human DAT and of other related SLC6 transporters.

Sensory evaluation and electronic tongue for sensing flavored mineral water taste attributes.

In this article a trained sensory panel evaluated 6 flavored mineral water samples. The samples consisted of 3 different brands, each with 2 flavors (pear-lemon grass and josta berry). The applied sensory method was profile analysis. Our aim was to analyze the sensory profiles and to investigate the similarities between the sensitivity of the trained human panel and an electronic tongue device. Another objective was to demonstrate the possibilities for the prediction of sensory attributes from electronic tongue measurements using a multivariate statistical method (Partial Least Squares regression [PLS]). The results showed that the products manufactured under different brand name but with the same aromas had very similar sensory profiles. The panel performance evaluation showed that it is appropriate (discrimination ability, repeatability, and panel consensus) to compare the panels results with the results of the electronic tongue. The samples can be discriminated by the electronic tongue and an accurate classification model can be built. Principal Component Analysis BiPlot diagrams showed that Brand A and B were similar because the manufacturers use the same aroma brands for their products. It can be concluded that Brand C was quite different compared to the other samples independently of the aroma content. Based on the electronic tongue results good prediction models can be obtained with high correlation coefficient (r(2) > 0.81) and low prediction error (RMSEP < 13.71 on the scale of the sensory evaluation from 0 to 100).

Sensory evaluation and electronic tongue analysis for sweetener recognition in coke drinks

Consumption of beverages with low energy has an increasing role. Furthermore hydrolyzed starch products such as inverted syrup show a wide application in the beverage industry. Therefore the importance of methods which can monitor the usage of natural and artificial sweeteners is increasing. The task was to describe the relevant sensory attributes and to determine the applicability of the electronic tongue to discriminate the coke drink samples with different sweeteners. Furthermore the aim was to find relationship between the taste attributes and measurement results provided by electronic tongue. An Alpha Astree Electronic Tongue and a trained sensory panel were used to evaluate the coke samples. Panelists found significant differences between the samples in 15 cases from the 18 sensory attributes defined previously by the consensus group. Coke drinks containing different kind of sweeteners can be characterized according to these sensory attributes. The samples were definitely distinguished by the electr...

Access path to the ligand binding pocket may play a role in xenobiotics selection by AhR

Understanding of multidrug binding at the atomic level would facilitate drug design and strategies to modulate drug metabolism, including drug transport, oxidation, and conjugation. Therefore we explored the mechanism of promiscuous binding of small molecules by studying the ligand binding domain, the PAS-B domain of the aryl hydrocarbon receptor (AhR). Because of the low sequence identities of PAS domains to be used for homology modeling, structural features of the widely employed HIF-2α and a more recent suitable template, CLOCK were compared. These structures were used to build AhR PAS-B homology models. We performed molecular dynamics simulations to characterize dynamic properties of the PAS-B domain and the generated conformational ensembles were employed in in silico docking. In order to understand structural and ligand binding features we compared the stability and dynamics of the promiscuous AhR PAS-B to other PAS domains exhibiting specific interactions or no ligand binding function. Our exhaustive in silico binding studies, in which we dock a wide spectrum of ligand molecules to the conformational ensembles, suggest that ligand specificity and selection may be determined not only by the PAS-B domain itself, but also by other parts of AhR and its protein interacting partners. We propose that ligand binding pocket and access channels leading to the pocket play equally important roles in discrimination of endogenous molecules and xenobiotics.

Sensing Basic Tastes by Electronic Tongue Sensors

There is an increasing demand to develop method for simulating the human taste perception by objective instruments1. The task was to develop method for the assessment of definite taste attributes. Therefore, our objective was to develop complete method for sensing different taste attributes. The subject of this work was to test the Specific Sensor Array for taste screening developed by Alpha M.O.S. Different brands of carrot juices were analyzed by an Alpha Astree Electronic Tongue (ET) and a trained sensory panel. The results of the sensory evaluation showed that the different carrot juice samples were significantly different from each other in some taste attributes. The electronic tongue was able to distinguish the tested samples according to the measurement results evaluated by multivariate statistics. Furthermore, the relevant taste attributes of carrot juice samples such as sour taste could be predicted by definite sensors of the electronic tongue. Based on our results we concluded that the selected ...

Taste attributes profiling in carrot juice using an electronic tongue

The taste of five brands of carrot juice was analyzed both by a sensory panel and an electronic tongue. The panelists found significant differences between the carrot juice samples in some appearance and odor attributes and in the relevant taste attributes such as sour taste, sweet taste and taste persistence. Principal component analysis plot calculated from the electronic tongue results showed a clear separation between the sample groups, with a ranking on sourness similar to the one from the panel.

Dopamine transporter oligomerization involves the scaffold domain, but spares the bundle domain

The human dopamine transporter (hDAT) is located on presynaptic neurons, where it plays an essential role in limiting dopaminergic signaling by temporarily curtailing high neurotransmitter concentration through rapid re-uptake. Transport by hDAT is energized by transmembrane ionic gradients. Dysfunction of this transporter leads to disease states, such as Parkinson’s disease, bipolar disorder or depression. It has been shown that hDAT and other members of the monoamine transporter family exist in oligomeric forms at the plasma membrane. Several residues are known to be involved in oligomerization, but interaction interfaces, oligomer orientation and the quarternary arrangement in the plasma membrane remain poorly understood. Here we examine oligomeric forms of hDAT using a direct approach, by following dimerization of two randomly-oriented hDAT transporters in 512 independent simulations, each being 2 μs in length. We employed the DAFT (docking assay for transmembrane components) approach, which is an unbiased molecular dynamics simulation method to identify oligomers, their conformations and populations. The overall ensemble of a total of >1 ms simulation time revealed a limited number of symmetric and asymmetric dimers. The identified dimer interfaces include all residues known to be involved in dimerization. Importantly, we find that the surface of the bundle domain is largely excluded from engaging in dimeric interfaces. Such an interaction would typically lead to inhibition by stabilization of one conformation, while substrate transport relies on a large scale rotation between the inward-facing and the outward-facing state.

Dissecting the Forces that Dominate Dimerization of the Nucleotide Binding Domains of ABCB1

P-glycoprotein, also known as multidrug resistance protein 1 or ABCB1, can export a wide range of chemically unrelated compounds, including chemotherapeutic drugs. ABCB1 consists of two transmembrane domains that form the substrate binding and translocation domain, and of two cytoplasmic nucleotide binding domains (NBDs) that energize substrate transport by ATP binding and hydrolysis. ATP binding triggers dimerization of the NBDs, which switches the transporter from an inward facing to an outward facing transmembrane domain conformation. We performed MD simulations to study the dynamic behavior of the NBD dimer in the presence or absence of nucleotides. In the apo configuration, the NBDs were overall attractive to each other as shown in the potential of mean force profile, but the energy well was shallow and broad. In contrast, a sharp and deep energy minimum (∼−42 kJ/mol) was found in the presence of ATP, leading to a well-defined conformation. Motif interaction network analyses revealed that ATP stabilizes the NBD dimer by serving as the central hub for interdomain connections. Simulations showed that forces promoting dimerization are multilayered, dominated by electrostatic interactions between the nucleotide and conserved amino acids of the signature sequence and the Walker A motif. In addition, direct and water-bridged hydrogen bonds between NBDs provided conformation-defining interactions. Importantly, we characterized a largely unrecognized but essential contribution from hydrophobic interactions between the adenine moiety of the nucleotides and a hydrophobic surface of the X-loop to the stabilization of the nucleotide-bound NBD dimer. These hydrophobic interactions lead to a sharp energy minimum, thereby conformationally restricting the nucleotide-bound state.