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Transporters of the solute carrier 6 (SLC6) family mediate the reuptake of neurotransmitters such as dopamine, norepinephrine, serotonin, GABA, and glycine. SLC6 family members are 12 transmembrane helix‐spanning proteins that operate using the transmembrane sodium gradient for transport. These transporters assume various quaternary arrangements ranging from monomers to complex stoichiometries with multiple subunits. Dopamine and serotonin transporter oligomerization has been implicated in trafficking of newly formed proteins from the endoplasmic reticulum to the plasma membrane with a pre‐fixed assembly. Once at the plasma membrane, oligomers are kept fixed in their quaternary assembly by interaction with phosphoinositides. While it remains unclear how oligomer formation precisely affects physiological transporter function, it has been shown that oligomerization supports the activity of release‐type psychostimulants. Most recently, single molecule microscopy experiments unveiled that the stoichiometry differs between individual members of the SLC6 family. The present overview summarizes our understanding of the influence of plasma membrane constituents on transporter oligomerization, describes the known interfaces between protomers and discusses open questions., We review the current knowledge on oligomerization of the solute carrier 6 (SLC6) family, which includes the transporters for the neurotransmitters dopamine, serotonin, and GABA. Early biochemical studies suggested dimer and tetramer configurations and identified several interaction motifs. Not all interfaces could be confirmed. Oligomers seem to become defined during transporter trafficking, are dependent on the plasma membrane PIP2 content, and affect substrate efflux. Simulations are starting to contribute to our understanding of transporter oligomerization, whereas recent single molecule experiments refined the oligomeric state by finding several non‐exchanging configurations for SERT, but only monomers and dimers for DAT.

The human serotonin transporter (hSERT) removes the neurotransmitter serotonin from the synaptic cleft by reuptake into the presynaptic nerve terminal. A number of neurologic diseases are associated with dysfunction of the hSERT, and several medications for their treatment are hSERT blockers, including citalopram, fluoxetine, and paroxetine. The substrate transport is energized by the high concentration of external NaCl. We showed through molecular dynamics simulations that the binding of NaCl stabilized the hSERT in the substrate-binding competent conformation, which was characterized by an open access path to the substrate-binding site through the outer vestibule. Importantly, the binding of NaCl reduced the dynamics of the hSERT by decreasing the internal fluctuations of the bundle domain as well as the movement of the bundle domain relative to the scaffold domain. In contrast, the presence of only the bound chloride ion did not reduce the high domain mobility of the apo state.

Derivatives of (2-aminopropyl)indole (API) and \ud (2-aminopropyl)benzofuran (APB) are new psychoactive substances which produce stimulant effects in vivo. \ud (2-Aminopropyl)benzo[β]thiophene (APBT) is a novel sulfur-based analog of API and APB that has not been pharmacologically characterized. In the current study, we assessed the pharmacological effects of six APBT positional isomers in vitro, and three of these isomers (3-APBT, 5-APBT, and 6-APBT) were subjected to further investigations in vivo. Uptake inhibition and efflux assays in human transporter-transfected HEK293 cells and in rat brain synaptosomes revealed that APBTs inhibit monoamine reuptake and induce transporter-mediated substrate release. Despite being non-selective transporter releasers like MDMA, the APBT compounds failed to produce locomotor stimulation in C57BL/6J mice. Interestingly, 3-APBT, 5-APBT, and 6-APBT were full agonists at 5-HT2 receptor subtypes as determined by calcium mobilization assays and induced the head-twitch response in C57BL/6J mice, suggesting psychedelic-like activity. Compared to their APB counterparts, ABPT compounds demonstrated that replacing the oxygen atom with sulfur results in enhanced releasing potency at the serotonin transporter and more potent and efficacious activity at 5-HT2 receptors, which fundamentally changed the in vitro and in vivo profile of APBT isomers in the present studies. Overall, our data suggest that APBT isomers may exhibit psychedelic and/or entactogenic effects in humans, with minimal psychomotor stimulation. Whether this unique pharmacological profile of APBT isomers translates into potential therapeutic potential, for instance as candidates for drug-assisted psychotherapy, warrants further investigation.

Structural data on ABCG5/G8 and ABCG2 reveal a unique molecular architecture for subfamily G ATP-binding cassette (ABCG) transporters and disclose putative substrate-binding sites. ABCG5/G8 and ABCG2 appear to use several unique structural motifs to execute transport, including the triple helical bundles, the membrane-embedded polar relay, the re-entry helices, and a hydrophobic valve. Interestingly, ABCG2 shows extreme substrate promiscuity, whereas ABCG5/G8 transports only sterol molecules. ABCG2 structures suggest a large internal cavity, serving as a binding region for substrates and inhibitors, while mutational and pharmacological analyses support the notion of multiple binding sites. By contrast, ABCG5/G8 shows a collapsed cavity of insufficient size to hold substrates. Indeed, mutational analyses indicate a sterol-binding site at the hydrophobic interface between the transporter and the lipid bilayer. In this review, we highlight key differences and similarities between ABCG2 and ABCG5/G8 structures. We further discuss the relevance of distinct and shared structural features in the context of their physiological functions. Finally, we elaborate on how ABCG2 and ABCG5/G8 could pave the way for studies on other ABCG transporters.

Temperature, memory effect, and cross-contamination are suspected to contribute to drift in electronic tongue (e-tongue) sensors, therefore drift corrections are required. This paper aimed to assess the disturbing effects on the sensor signals during measurement with an Alpha Astree e-tongue and to develop drift correction techniques. Apple juice samples were measured at different temperatures. pH change of apple juice samples was measured to assess cross-contamination. Different sequential orders of model solutions and apple juice samples were applied to evaluate the memory effect. Model solutions corresponding to basic tastes and commercial apple juice samples were measured for six consecutive weeks to model drift of the sensor signals. Result showed that temperature, cross-contamination, and memory effect influenced the sensor signals. Three drift correction methods: additive drift correction based on all samples, additive drift correction based on reference samples, and multi sensor linear correction, were developed and compared to the component correction in literature through linear discriminant analysis (LDA). LDA analysis showed all the four methods were effective in reducing sensor drift in long-term measurements but the additive correction relative to the whole sample set gave the best results. The results could be explored for long-term measurements with the e-tongue.

The human ATP-binding cassette transporter ABCG2 is a key to anticancer resistance and physiological detoxification. However, the molecular mechanism of substrate transport remains enigmatic. A hydrophobic di-leucine motif in the ABCG2 core separates a large intracellular cavity from a smaller upper cavity. We show that the di-leucine motif acts as a valve that controls drug extrusion. Moreover, the extracellular structure engages the re-entry helix and all extracellular loops to form a roof architecture on top of the upper cavity. Disulfide bridges and a salt bridge limit roof flexibility, but provide a lid-like function to control drug release. We propose that drug translocation from the central to the upper cavities through the valve is driven by a squeezing motion, suggesting that ABCG2 operates similar to a peristaltic pump. Finally, the roof contains essential residues, offering therapeutic options to block ABCG2 by either targeting the valve or essential residues in the roof., The human ATP-binding cassette transporter ABCG2 plays critical roles in anticancer resistance but the molecular mechanism of ABCG2-mediated substrate transport remains enigmatic. Here authors use extensive mutagenesis and molecular dynamics simulations to reveal a mechanistic basis for the function of the di-leucine valve and the roof organization in the transport cycle.

Neurotransmitter:sodium symporters are highly expressed in the human brain and catalyze the uptake of substrate through the plasma membrane by using the electrochemical gradient of sodium as the energy source. The bacterial homolog LeuT, a small amino acid transporter isolated from the bacteria Aquifex aeolicus, is the founding member of the family and has been crystallized in three conformations. The N-terminus is structurally well defined and strongly interacts with the transporter core in the outward-facing conformations. However, it could not be resolved in the inward-facing conformation, which indicates enhanced mobility. Here we investigate conformations and dynamics of the N-terminus, by combining molecular dynamics simulations with experimental verification using distance measurements and accessibility studies. We found strongly increased dynamics of the N-terminus, but also that helix TM1A is subject to enhanced mobility. TM1A moves towards the transporter core in the membrane environment, reaching a conformation that is closer to the structure of LeuT with wild type sequence, indicating that the mutation introduced to create the inward-facing structure might have altered the position of helix TM1A. The mobile N-terminus avoids entering the open vestibule of the inward-facing state, as accessibility studies do not show any reduction of quenching by iodide of a fluorophore attached to the N-terminus.

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.

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., Author summary The human dopamine transporter efficiently removes the neurotransmitter dopamine from the synaptic cleft. Alteration of dopamine transporter function is associated with several neurological diseases, including mood disorders or attention-deficit hyperactivity disorder, but is also a major player in addiction and drug abuse. Functional studies have revealed that not only is transporter oligomerization involved in surface expression and endocytosis, but, more importantly, in reverse transport (efflux) of dopamine that is triggered by amphetamine-like drugs of abuse. Structural knowledge of transporter oligomerization is largely missing. We performed a large scale comprehensive computational study on transporter oligomerization to reveal dimer geometries and the residues involved in the interfaces. The dimer conformations we find in our dataset are fully consistent with all available experimental data in the literature, but also show novel interfaces. We further verified all dimer geometries by free energy calculations. Our results identified an unpredicted—but for the mechanism of substrate transport essential—property: the bundle domain, which moves during the transport cycle, is excluded from contributing to dimer interfaces, thereby allowing for unrestrained movements necessary to translocate substrates through the membrane.

ABC (ATP binding cassette) transporters, ubiquitous in all kingdoms of life, carry out essential substrate transport reactions across cell membranes. Their transmembrane domains bind and translocate substrates and are connected to a pair of nucleotide binding domains, which bind and hydrolyze ATP to energize import or export of substrates. Over four decades of investigations into ABC transporters have revealed numerous details from atomic-level structural insights to their functional and physiological roles. Despite all these advances, a comprehensive understanding of the mechanistic principles of ABC transporter function remains elusive. The human multidrug resistance transporter ABCB1, also referred to as P-glycoprotein (P-gp), is one of the most intensively studied ABC exporters. Using ABCB1 as the reference point, we aim to compare the dominating mechanistic models of substrate transport and ATP hydrolysis for ABC exporters and to highlight the experimental and computational evidence in their support. In particular, we point out in silico studies that enhance and complement available biochemical data. “This article is part of a Special Issue entitled: Beyond the Structure Function Horizon of Membrane Proteins edited by Ute Hellmich, Rupak Doshi and Benjamin McIlwain.”

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.

Several efforts have been made by researchers to predict the sensory profile of coffee by instrumental measurement results. The aim of the work reported here was to determine the applicability of instrumental methods on coffee samples to replace the sensory analysis. For this purpose, we evaluated the most important sensory attributes of coffee capsule samples by sensory evaluation, electronic tongue and chemical analysis (SPME–GC–MS). The trained sensory panel was able to distinguish the samples based on global odor (p = 0.01), and bitter (p

Nowadays quality measurement is an important topic in food quality control. The electronic tongue (ET) can be a useful tool in this feld. The objective of our work is to demonstrate the application potential of ET for the evaluation of different coffee, wine and carrot juice samples and to compare the results with sensory attributes. ET was able to distinguish the different coffee samples. The Arabica concentration of the samples were predicted with close correlation (R2=0.98) and low error (RMSEP=3.16). The Arabica content of commercial samples were also determined. The ET measurement results of different wine samples showed a tendency similar to the increasing ‘acidic content’ determined by sensory evaluation. The closest correlation between ET and sensory evaluation was found with the ‘acidic taste’ (R2=0.87) and the lowest prediction error was observed with the prediction of ’fruit taste’ (RMSEP=6.11). Carrot juice samples were also distinguished by ET. Sensor SRS, developed for sour taste, gave the highest correlation (r=−0.99) with the sour taste of the carrot juice samples. The conclusion is that ET is a useful instrument in the feld of food quality control when appropriate statistical methods are applied.

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 panel's 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).

Efforts have been made to predict the sensory profile of coffee samples by instrumental measurement results. The objective of the work was to evaluate the most important sensory attributes of coffee samples prepared from ground roasted coffee by electronic tongue and by sensory panel. Further aim was to predict the Arabica concentration and the main sensory attributes of the different coffee blends by electronic tongue and to analyze the sensitivity of the electronic tongue to the detection of poor quality coffee samples. Five coffee blends with known Arabica and Robusta concentration ratio, five commercially available coffee blends and a poor quality coffee were analyzed. The electronic tongue distinguished the coffee samples according to the Arabica and Robusta content. The sensory panel was able to discriminate the samples based on global aroma, bitterness and coffee aroma intensity (p < 0.01). The Arabica concentration was predicted from the electronic tongue results by PLS with close correlation and low prediction error. Models were developed to predict sensory attributes of the tested coffee samples from the results obtained by the electronic instrument.

ABC (ATP Binding Cassette) proteins form one of the largest protein superfamilies. Most members are active membrane transporters translocating their substrates across the lipid bilayer of the plasma membrane or intracellular organelles. Multidrug transporters exhibit broad substrate specificity, exporting molecules with diverse chemical structures to protect organisms from xenotoxic compounds, and also play an important role in influencing the efficacy of therapeutic agents. High resolution structural information is required to reveal the conformational changes associated with the transport cycle and the interaction with small molecules, with the ultimate aim to develop strategies to pharmacologically modulate function and predict substrates properties. In this chapter we review available ABC protein structures and discuss advances in using this structural information for computational approaches that are aimed at elucidating the mechanism of substrate recognition and cargo translocation in the context of the ATP catalytic cycle of human multidrug ABC transporters.

Munkamban az Alpha Astree elektronikus nyelv meresi eredmenyeit befolyasolo iz kolcsonhatasokat igyekeztem felderiteni a hatasok globalis es szenzor szintű elemzesevel. Az elektronikus nyelvet erintő mintaosszetevő kolcsonhatasok vizsgalata jelentősen hozzajarul az elektronikus nyelvről szerzett elmeleti tudasunkhoz. Ezen tulmenően hasznos gyakorlati javaslatokkal szolgal mind kiserlettervezesi, mind kiertekelesi szempontbol. A kolcsonhatasok vizsgalatahoz az ot szabvanyosnak tekinthető, alapizeknek megfelelő vegyuletből keszitett tiszta es kombinalt oldatokat es elelmiszer mintakat vizsgaltam egy, az ismert zavaro hatasokat minimalizalo meresi modszerrel.

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), Cohen's 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 © 2012 John Wiley & Sons, Ltd.

Reliability and validity of sensory data is an important issue in scientific researches. If sensory analysis is performed in an analytical approach, the resulting data will show a similar structure to the chemical analyses. In the present paper the authors have used a complex approach to evaluate the performance of a sensory panel. The tested samples were black tea batches from different plantations of Sri Lanka. Profile analysis was applied to identify the odor profiles of the samples. Sensory profile data was submitted to two novel techniques of panel performance evaluation. GCAP (Gravity Center Area/Perimeter) is based on the profile polygons of the individual assessors. If the area/perimeter ratio of two panelists' profiles is similar and the gravity center is located near to each other, the panelists performed the tests consistently. CRRN (Compare Ranks with Random Numbers) is applicable not only to sensory data but also to other field of chemometrics. The essence of CRRN method is based on the evaluation of an ‘average’ vector, corresponding to the coordinate-averages of the measured points, and on a produced random vector series of the same dimension as the measured points. Sensory and e-nose data were evaluated with principal component analysis, cluster analysis and linear discriminant analysis. Partial least square regression and support vector machine regression were used to predict sensory data with electronic nose results. Prediction by support vector machine gave close correlation between the results of electronic nose measurement and odor attributes. Copyright © 2011 John Wiley & Sons, Ltd.

The objective of the work reported was to predict some sensory attributes of carrots stored under non-ideal conditions from the data obtained on taste measured by electronic tongue and on the physical properties (acoustic stiffness, cutting force, deformation work ratio and luminosity). There was a close correlation between the mechanical characteristics and the non-ideal storage time. Sensory evaluation showed significant ranking in “bite and chewing”, “sweet taste” and “global impression” attributes according to the Page test. Principal component analysis (PCA) plots were determined for the acoustic stiffness coefficient, cutting force and deformation work ratio and these showed that PC1 followed a tendency similar to that of the storage time. PCA plots were determined for the electronic tongue measurements and this PCA separated the sample groups along PC1 and PC2. We used partial least square (PLS) regression to predict “bite and chewing” from the acoustic stiffness coefficient, cutting force, and def...

The objective of the research was to compare the taste attributes of different commercial soya drinks. Furthermore, the task was to determine the effect of different ingredients and processing technologies on the taste attributes of the product. Based on the results of electronic tongue measurements the instrument is able to determine the effect of the applied technology and to distinguish soya juice samples according to sensory preferences. Canonical discriminant analysis showed that the groups of two measurements of the same products were overlapping. Therefore, the electronic tongue measurements are supposed to be of acceptable repeatability. The canonical discriminant analysis showed that the taste attributes of soya juice made of hulled soybeans was beneficial for the taste attributes relative to that of the juice made of not-hulled soybeans. Three main groups could be observed from the analyzed six commercial soya drink samples based on canonical discriminant analysis. There is a group of top market brands having definite taste improver additives and another one containing three products having low amount of additives. However, the group of samples made of soybean and rice is located between the above-mentioned two groups in the discriminant score plot.

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.

The quality of fruit based spirits varies year to year; therefore, the identification of the vintage of a distilled alcoholic beverage is necessary, but requires highly sensitive analytics. The interpretation of the gathered data requires a well-adapted chemometric method. In this study, Hungarian apple, sour cherry and plum distillates (pálinka’s) from different vintages were analyzed, classified and identified using volatile composition analyzed by GC-MS. The fruit’s origin, fermentation technique and distillation were the same at all the fruits; the only differences in the samples were their vintages (2010, 2011 and 2012). Analysis of variance (ANOVA) and Linear discriminant analysis (LDA) was applied for classification and components’ identification related to the vintage effect. The samples were successfully classified (correct classification rate ranging from 75 to 100%), three components are found to be related to the vintage effect regardless the fruit type: propanol, butanol and ethyl-propionate. GC-MS data proved to be a promising tool for classification of fruit distillate vintages.

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 panel's 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 (RMSEP13.71 on the scale of the sensory evaluation from 0 to 100).

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 electronic tongue. The main difference was found between the samples made with natural and artificial sweeteners. However electronic tongue was able to distinguish samples containing different kind of artificial and different kind of natural sweeteners, as well. Taste attributes of coke drinks determined by sensory panel were predicted by partial least squares regression method based on the results of electronic tongue with close correlation and low prediction error.

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 sensors of the Specific Sensor Array could be an appropriate tool for estimating important taste attributes of the tested carrot juice samples.

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.

The plasmalemmal norepinephrine transporter (NET) regulates cardiovascular sympathetic activity by clearing extracellular norepinephrine in the synaptic cleft. Here, we investigate the subunit stoichiometry and function of NET using single-molecule fluorescence microscopy and flux assays. In particular, we show the effect of phosphatidylinositol 4,5-bisphosphate (PIP2) on NET oligomerization and efflux. NET forms monomers (~60%) and dimers (~40%) at the plasma membrane. PIP2 depletion results in a decrease in the average oligomeric state and decreases NET-mediated substrate efflux while not affecting substrate uptake. Mutation of the putative PIP2 binding residues R121, K334, and R440 to alanines does not affect NET dimerization but results in decreased substrate efflux that is not altered upon PIP2 depletion; this indicates that PIP2 interactions with these residues affect NET-mediated efflux. A dysregulation of norepinephrine and PIP2 signaling have both been implicated in neuropsychiatric and cardiovascular diseases. This study provides evidence that PIP2 directly regulates NET organization and function.

The human serotonin transporter (hSERT) terminates neurotransmission by removing serotonin (5HT) from the synaptic cleft, an essential process for proper functioning of serotonergic neurons. Structures of the hSERT have revealed its molecular architecture in four conformations, including the outward-open and occluded states, and show the transporter's engagement with co-transported ions and the binding mode of inhibitors. In this study, we investigated the molecular mechanism by which the hSERT occludes and sequesters the substrate 5HT. This first step of substrate uptake into cells is a structural change consisting of the transition from the outward-open to the occluded state. Inhibitors such as the antidepressants citalopram, fluoxetine, and sertraline inhibit this step of the transport cycle. Using molecular dynamics simulations, we reached a fully occluded state, in which the transporter-bound 5HT becomes fully shielded from both sides of the membrane by two closed hydrophobic gates. Analysis of 5HT-triggered occlusion showed that bound 5HT serves as an essential trigger for transporter occlusion. Moreover, simulations revealed a complex sequence of steps and showed that movements of bundle domain helices are only partially correlated. 5HT-triggered occlusion is initially dominated by movements of transmembrane helix 1b, while in the final step, only transmembrane helix 6a moves and relaxes an intermediate change in its secondary structure.