Catalysis driven by an amyloid-substrate complex.

Amine modification through nucleophilic attack of the amine functionality is a very common chemical transformation. Under biorelevant conditions using acidic-to-neutral pH buffer, however, the nucleophilic reaction of alkyl amines (pKa ≈ 10) is not facile due to the generation of ammonium ions lacking nucleophilicity. Here, we disclose a unique molecular transformation system, catalysis driven by amyloid-substrate complex (CASL), that promotes amine modifications in acidic buffer. Ammonium ions attached to molecules with amyloid-binding capability were activated through deprotonation due to the close proximity to the amyloid catalyst formed by Ac-Asn-Phe-Gly-Ala-Ile-Leu-NH2 (NL6), derived from islet amyloid polypeptide (IAPP). Under the CASL conditions, alkyl amines underwent various modifications, i.e., acylation, arylation, cyclization, and alkylation, in acidic buffer. Crystallographic analysis and chemical modification studies of the amyloid catalysts suggested that the carbonyl oxygen of the Phe-Gly amide bond of NL6 plays a key role in activating the substrate amine by forming a hydrogen bond. Using CASL, selective conversion of substrates possessing equivalently reactive amine functionalities was achieved in catalytic reactions using amyloids. CASL provides a unique method for applying nucleophilic conversion reactions of amines in diverse fields of chemistry and biology.

Insight into chemically reactive metabolites of aliphatic amine pollutants: A de novo prediction strategy and case study of sertraline.

The uncommon metabolic pathways of organic pollutants are easily overlooked, potentially leading to idiosyncratic toxicity. Prediction of their biotransformation associated with the toxic effects is the very purpose that this work focuses, to develop a de novo method to mechanistically predict the reactive toxicity pathways of uncommon metabolites from start aliphatic amine molecules, which employed sertraline triggered by CYP450 enzymes as a model system, as there are growing concerns about the effects on human health posed by antidepressants in the aquatic environment. This de novo prediction strategy combines computational and experimental methods, involving DFT calculations upon sequential growth, in vitro and in vivo assays, dissecting chemically reactive mechanism relevant to toxicity, and rationalizing the fundamental factors. Significantly, desaturation and debenzylation-aromatization as the emerging metabolic pathways of sertraline have been elucidated, with the detection of DNA adducts of oxaziridine metabolite in mice, highlighting the potential reactive toxicity. Molecular orbital analysis supports the reactivity preference for toxicological-relevant C-N desaturation over N-hydroxylation of sertraline, possibly extended to several other aliphatic amines based on the Bell-Evans-Polanyi principle. It was further validated toward some other wide-concerned aliphatic amine pollutants involving atrazine, ε-caprolactam, 6PPD via in silico and in vitro assays, thereby constituting a complete path for de novo prediction from case study to general applications.

Measuring bioavailability, utilization, and excretion of rumen-protected lysine in lactating cows using an isotope technique.

Supplementing a diet with rumen-protected amino acids (AAs) is a common feeding strategy for efficient production. For a cost-effective use of rumen-protected AA, the accurate bioavailability of rumen-protected amino acids should be known and their metabolism after absorption needs to be well understood. The current study determined the bioavailability, absorption, utilization, and excretion of rumen-protected Lys (RP-Lys). Four ruminally cannulated cows in a 4 × 4 Latin square design (12 d for diet adaptation; 5 or 6 d for total collections) received the following treatments: L0, a basal diet; L25, the basal diet and L-Lys infused into the abomasum to provide 25.9 g/d L-Lys; L50, the basal diet and L-Lys infused into the abomasum to provide 51.8 g/d L-Lys; and RPL, the basal diet supplemented with 105 g/d (as-is) of RP-Lys to provide 26.7 g of digestible Lys. During the last 5 or 6 d in each period, 15N-Lys (0.38 g/d) was infused into the abomasum for all cows to label the pool of AA, and the total collection of milk, urine, and feces were conducted. 15N enrichment of samples on d 4 and 5 were used to calculate the bioavailability and Lys metabolism. We used a model containing a fast AA turnover (≤ 5 d) and slow AA turnover pool (> 5 d) to calculate fluxes of Lys. The Lys flux to the fast AA turnover pool (absorbed Lys + Lys from the slow AA turnover pool to fast AA turnover pool) was calculated using 15N enrichment of milk Lys. The flux of Lys from the fast AA turnover pool to milk and urine was calculated using 15N transfer into milk and urine. Then, absorbed Lys was estimated by the sum of Lys flux to milk and urine assuming no net utilization of Lys by body tissues. Duodenal Lys flow was estimated by 15N enrichment of fecal Lys. The bioavailability of RP-Lys was calculated from duodenal Lys flows and Lys absorption for RPL. Increasing Lys supply from L25 to L50 increased Lys utilization for milk by 9 g/d but also increased urinary excretion by 10 g/d. For RPL, absorbed Lys was estimated to be 136 g/d where 28 g of absorbed Lys originated from RP-Lys. In conclusion, 68% of bioavailability was obtained for RP-Lys. The Lys provided from RP-Lys was not only utilized for milk protein (48%) but also excreted in urine (20%) after oxidation.

The effect of valproate on the amino acids, monoamines, and kynurenic acid concentrations in brain structures involved in epileptogenesis in the pentylenetetrazol-kindled rats.

BACKGROUND: The study aimed to assess the influence of a single valproate (VPA) administration on inhibitory and excitatory neurotransmitter concentrations in the brain structures involved in epileptogenesis in pentylenetetrazol (PTZ)-kindled rats. METHODS: Adult, male Wistar rats were kindled by repeated intraperitoneal (ip) injections of PTZ at a subconvulsive dose (30 mg/kg, three times a week). Due to the different times required to kindle the rats (18-22 injections of PTZ), a booster dose of PTZ was administrated 7 days after the last rats were kindled. Then rats were divided into two groups: acute administration of VPA (400 mg/kg) or saline given ip. The concentration of amino acids, kynurenic acid (KYNA), monoamines, and their metabolites in the prefrontal cortex, hippocampus, amygdala, and striatum was assessed by high-pressure liquid chromatography (HPLC). RESULTS: It was found that a single administration of VPA increased the gamma-aminobutyric acid (GABA), tryptophan (TRP), 5-hydroxyindoleacetic acid (5-HIAA), and KYNA concentrations and decreased aspartate (ASP) levels in PTZ-kindled rats in the prefrontal cortex, hippocampus, amygdala and striatum. CONCLUSIONS: Our results indicate that a single administration of VPA in the PTZ-kindled rats restored proper balance between excitatory (decreasing the level of ASP) and inhibitory neurotransmission (increased concentration GABA, KYNA) and affecting serotoninergic neurotransmission in the prefrontal cortex, hippocampus, amygdala, and striatum.

High Cell Density Cultivation Combined with high Specific Enzyme Activity: Cultivation Protocol for the Production of an Amine Transaminase from Bacillus megaterium in E. coli.

High cell density cultivation is an established method for the production of various industrially important products such as recombinant proteins. However, these protocols are not always suitable for biocatalytic processes as the focus often lies on biomass production rather than high specific activities of the enzyme inside the cells. In contrast, a range of shake flask protocols are well known with high specific activities but rather low cell densities. To overcome this gap, we established a tailor-made fed-batch protocol combining both aspects: high cell density and high specific activities of heterologously produced enzyme. Using the example of an industrially relevant amine transaminase from Bacillus megaterium, we describe a strategy to optimize the cultivation yield based on the feed rate, IPTG concentration, and post-induction temperature. By adjusting these key parameters, we were able to increase the specific activity by 2.6-fold and the wet cell weight by even 17-fold compared to shake flasks. Finally, we were able to verify our established protocol by transferring it to another experimenter. With that, our optimization strategy can serve as a template for the production of high titers of heterologously produced, active enzymes and might enable the availability of these catalysts for upscaling biocatalytic processes.

Bile salt hydrolase catalyses formation of amine-conjugated bile acids.

Bacteria in the gastrointestinal tract produce amino acid bile acid amidates that can affect host-mediated metabolic processes1-6; however, the bacterial gene(s) responsible for their production remain unknown. Herein, we report that bile salt hydrolase (BSH) possesses dual functions in bile acid metabolism. Specifically, we identified a previously unknown role for BSH as an amine N-acyltransferase that conjugates amines to bile acids, thus forming bacterial bile acid amidates (BBAAs). To characterize this amine N-acyltransferase BSH activity, we used pharmacological inhibition of BSH, heterologous expression of bsh and mutants in Escherichia coli and bsh knockout and complementation in Bacteroides fragilis to demonstrate that BSH generates BBAAs. We further show in a human infant cohort that BBAA production is positively correlated with the colonization of bsh-expressing bacteria. Lastly, we report that in cell culture models, BBAAs activate host ligand-activated transcription factors including the pregnane X receptor and the aryl hydrocarbon receptor. These findings enhance our understanding of how gut bacteria, through the promiscuous actions of BSH, have a significant role in regulating the bile acid metabolic network.

Calcium imaging of adult olfactory epithelium reveals amines as important odor class in fish.

The odor space of aquatic organisms is by necessity quite different from that of air-breathing animals. The recognized odor classes in teleost fish include amino acids, bile acids, reproductive hormones, nucleotides, and a limited number of polyamines. Conversely, a significant portion of the fish olfactory receptor repertoire is composed of trace amine-associated receptors, generally assumed to be responsible for detecting amines. Zebrafish possess over one hundred of these receptors, but the responses of olfactory sensory neurons to amines have not been known so far. Here we examined odor responses of zebrafish olfactory epithelial explants at the cellular level, employing calcium imaging. We report that amines elicit strong responses in olfactory sensory neurons, with a time course characteristically different from that of ATP-responsive (basal) cells. A quantitative analysis of the laminar height distribution shows amine-responsive cells undistinguishable from ciliated neurons positive for olfactory marker protein. This distribution is significantly different from those measured for microvillous neurons positive for transient receptor potential channel 2 and basal cells positive for proliferating cell nuclear antigen. Our results suggest amines as an important odor class for teleost fish.

[Effect of a novel pH-responsive tertiary amine monomer dodecylmethylaminoethyl methacrylate modified resin adhesive on biofilm formation of Streptococcus mutans and Lactobacillus casei in vitro].

Objective: To explore the application prospect of a new pH-responsive tertiary amine monomer dodecylmethylaminoethyl methacrylate (DMAEM) modified resin adhesive (DMAEM@RA) in the prevention and treatment of secondary caries. Methods: Five percents DMAEM was added to the resin adhesive to synthesize DMAEM@RA for modifying. Streptococcus mutans (Sm) and Lactobacillus casei (Lc) biofilms were cultured on resin adhesive and DMAEM@RA, respectively. The culture systems were set up at pH=7.4, 6.0, 5.5, and 5.0. The antimicrobial activity of DMAEM@RA was evaluated by quantitative PCR. The effects of DMAEM@RA on biofilm thickness, bacterial amount, and extracellular polysaccharides were studied by scanning electron microscope (SEM) and extracellular polysaccharide staining. Real-time fluorescence quantitative PCR was used to study the effect of DMAEM@RA on the expression levels of cariogenic genes in Sm. Results: DMAEM@RA could significantly reduce the amount of Sm and Lc under acidic conditions, especially Lc. At pH=5.0, the logarithm value of co-cultured Sm bacteria [lg (CFU/ml)] in DMAEM@RA group (7.58±0.01) was significantly lower than that in control group (7.87±0.03) (t=14.32, P<0.001), and the logarithm value of Lc bacteria [lg (CFU/ml)] (7.29±0.04) was also significantly lower than that in control group (7.93±0.15) (t=6.93, P=0.002). SEM observed that the bacteria decreased and the cell fragments appeared in DMAEM@RA group. In addition, DMAEM@RA significantly reduced the biomass of extracellular polysaccharides in the dual-species biofilm under acidic conditions. At pH=5.0, the biomass of extracellular polysaccharides in DMAEM@RA group [(25.13±3.14) mm3/mm2] was significantly lower than that in the control group [(42.66±7.46) mm3/mm2] (t=3.75, P=0.020). DMAEM@RA could significantly up-regulate the expressions of gtfB and gtfC genes in Sm under acidic conditions. At pH=5.0, gtfB and gtfC genes were significantly up-regulated by (14.64± 0.44) times and (2.99±0.20) times, respectively (t=-42.74, P<0.001; t=-13.55, P<0.001). Conclusions: The DMAEM@RA has a good antibacterial effect under acidic conditions, demonstrating that it has a good potential to prevent the occurrence and development of secondary caries.

Radiotherapy-activated NBTXR3 nanoparticles promote ferroptosis through induction of lysosomal membrane permeabilization.

PURPOSE: Radiotherapy-activated NBTXR3 (NBTXR3 + RT) has demonstrated superior efficacy in cancer cell destruction and tumor growth control, compared to radiotherapy (RT), in preclinical and clinical settings. Previous studies highlighted the immunomodulatory properties of NBTXR3 + RT, such as modification of tumor cell immunogenicity/adjuvanticity, producing an effective local tumor control and abscopal effect, related to an enhanced antitumor immune response. Furthermore, NBTXR3 + RT has shown potential in restoring anti-PD1 efficacy in a refractory tumor model. However, the early events leading to these results, such as NBTXR3 endocytosis, intracellular trafficking and primary biological responses induced by NBTXR3 + RT remain poorly understood. METHODS: We analyzed by transmission electron microscopy endocytosis and intracellular localization of NBTXR3 nanoparticles after endocytosis in various cell lines, in vitro and in vivo. A kinetic of NBTXR3 endocytosis and its impact on lysosomes was conducted using LysoTracker staining, and a RNAseq analysis was performed. We investigated the ability of NBTXR3 + RT to induce lysosomal membrane permeabilization (LMP) and ferroptosis by analyzing lipid peroxidation. Additionally, we evaluated the recapture by cancer cells of NBTXR3 released from dead cells. RESULTS: NBTXR3 nanoparticles were rapidly internalized by cells mainly through macropinocytosis and in a less extend by clathrin-dependent endocytosis. NBTXR3-containing endosomes were then fused with lysosomes. The day following NBTXR3 addition, we measured a significant increase in LysoTracker lysosome labeling intensity, in vitro as in vivo. Following RT, a significant lysosomal membrane permeabilization (LMP) was measured exclusively in cells treated with NBTXR3 + RT, while RT had no effect. The day post-irradiation, a significant increase in lipid peroxidation, a biomarker of ferroptosis, was measured with NBTXR3 + RT compared to RT. Moreover, we demonstrated that NBTXR3 nanoparticles released from dead cells can be recaptured by cancer cells. CONCLUSIONS: Our findings provide novel insights into the early and specific biological effects induced by NBTXR3 + RT, especially LMP, not induced by RT in our models. The subsequent significant increase in lipid peroxidation partially explains the enhanced cancer cell killing capacity of NBTXR3 + RT compared to RT, potentially by promoting ferroptosis. This study improves our understanding of the cellular mechanisms underlying NBTXR3 + RT and highlights its potential as an agnostic therapeutic strategy for solid cancers treatment.

Discovery and biocatalytic characterization of opine dehydrogenases by metagenome mining.

Enzymatic processes play an increasing role in synthetic organic chemistry which requires the access to a broad and diverse set of enzymes. Metagenome mining is a valuable and efficient way to discover novel enzymes with unique properties for biotechnological applications. Here, we report the discovery and biocatalytic characterization of six novel metagenomic opine dehydrogenases from a hot spring environment (mODHs) (EC 1.5.1.X). These enzymes catalyze the asymmetric reductive amination between an amino acid and a keto acid resulting in opines which have defined biochemical roles and represent promising building blocks for pharmaceutical applications. The newly identified enzymes exhibit unique substrate specificity and higher thermostability compared to known examples. The feature that they preferably utilize negatively charged polar amino acids is so far unprecedented for opine dehydrogenases. We have identified two spatially correlated positions in their active sites that govern this substrate specificity and demonstrated a switch of substrate preference by site-directed mutagenesis. While they still suffer from a relatively narrow substrate scope, their enhanced thermostability and the orthogonality of their substrate preference make them a valuable addition to the toolbox of enzymes for reductive aminations. Importantly, enzymatic reductive aminations with highly polar amines are very rare in the literature. Thus, the preparative-scale enzymatic production, purification, and characterization of three highly functionalized chiral secondary amines lend a special significance to our work in filling this gap. KEY POINTS: • Six new opine dehydrogenases have been discovered from a hot spring metagenome • The newly identified enzymes display a unique substrate scope • Substrate specificity is governed by two correlated active-site residues.

Amino acid standardized ileal digestibility together with concentrations of digestible and metabolizable energy in Saccharomyces cerevisiae yeast and soybean meal for gestating sows.

The standardized ileal digestibility (SID) of amino acids (AAs) plus crude protein (CP), in addition to digestible energy (DE) and metabolizable energy (ME) concentrations, was assessed through two experiments on Saccharomyces cerevisiae yeast (SCY) combined with soybean meal (SBM) for gestating sows. SCY and SBM were subjected to experiment 1 for the determination of CP and AAs in terms of SID. Under a randomized complete block design, three dietary treatments were provided for a total of 24 Landrace × Yorkshire gestating sows (parity 2), with the distal ileum clipped by a T-cannula at gestational day 33 based on body weight (BW) (194.1 ±â€…7.1, 195.3 ±â€…8.5, and 195.3 ±â€…8.6 kg). SCY and SBM were used as the only source of nitrogen to prepare two semi-purified diets and a nitrogen-free diet was also utilized to examine CP plus AAs for basal ileal endogenous losses. The gestating sows were initially fed these diets for 5 d to allow for adaptation, and ileal digesta was collected 2 d later for analysis. CP and all AAs in SCY, except for Trp and Gly, showed significantly lower SID than those in SBM (P < 0.05). Among the essential AAs, the range of SID was 68.8% for Thr to 92.2% for Arg in dried yeast, and from 79.9% for Thr to 98.6% for Met in SBM. DE plus ME were measured via experiment 2 with a randomized complete block design on SCY and SBM. Eighteen day-35 Landrace × Yorkshire pregnant sows (parity 3) were allocated to three diets based on BW (233.3 ±â€…16.0, 233.4 ±â€…9.6, and 233.4 ±â€…10.3 kg). Three diets were adopted for the experiment, namely, a corn-based diet as well as two diets containing 20.2% SCY and 20.0% SBM samples. The full fecal collection method, comprising a 5-day adaptation period before a 5- to 6-d experimental period for quantitative urine and feces collection, was employed for metabolic trials. The DE and ME for SCY were remarkably decreased compared with those for SBM (3812 kcal/kg DM vs. 4264 kcal/kg DM and 3714 kcal/kg DM vs. 4157 kcal/kg DM), respectively (P < 0.05). No differences were observed in the apparent total tract digestibility (ATTD) of organic matter, CP, and gross energy between SCY and SBM, but ATTD was significantly reduced in SCY for acid detergent fiber, dry matter, and neutral detergent fiber by contrast with SBM (P < 0.05). In conclusion, most AAs and CP in SCY had lower SID, DE, and ME than SBM in this study. These findings can be applied to diet formulation with the aforementioned ingredients for sows.

Saccharomyces cerevisiae yeast (SCY) is commonly used as an additive in feed (<1% of the formulation), but there is a limited amount of available information about its function as a promising source of proteins for pig diets, and especially, the nutritive value of yeast protein for gestating sows remains to be clarified. Feeding stuff has different digestibility between growing and gestating pigs. Therefore, our study evaluated the standardized ileal digestibility of amino acids and crude protein together with metabolizable and digestible energy in SCY for gestating sows, to provide nutritional value parameters on its potential as an effective alternative to traditional protein ingredients such as soybean meal in sow diet formulations.

Amino Acids Fueling Fibroblast-Like Synoviocyte Activation and Arthritis By Regulating Chemokine Expression and Leukocyte Migration.

OBJECTIVE: We analyzed the impact of amino acid (AA) availability on the inflammatory response in arthritis. METHODS: We stimulated rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLSs) with tumor necrosis factor (TNF) in the presence or absence of proteinogenic AAs and measured their response by QuantSeq 3' messenger RNA sequencing, quantitative polymerase chain reaction, and enzyme-linked immunosorbent assay. Signal transduction events were determined by Western blot. We performed K/BxN serum transfer arthritis in mice receiving a normal and a low-protein diet and analyzed arthritis clinically and histologically. RESULTS: Deprivation of AAs decreased the expression of a specific subset of genes, including the chemokines CXCL10, CCL2, and CCL5 in TNF-stimulated FLSs. Mechanistically, the presence of AAs was required for the TNF-induced activation of an interferon regulatory factor 1 (IRF1)-STAT1 signaling circuit that drives the expression of chemotactic factors. The expression of IRF1 and the IRF1-dependent gene set in FLSs was highly correlated with the presence of inflammatory cells in human RA, emphasizing the important role of this AA-dependent pathway in inflammatory cell recruitment to the synovial tissue. Finally, we show that mice receiving a low-protein diet expressed less IRF1 in the inflamed synovium and consequently developed reduced clinical and histologic signs of arthritis. CONCLUSION: AA deprivation reduces the severity of arthritis by suppressing the expression of IRF1-STAT1-driven chemokines, which are crucial for leukocyte recruitment to the arthritic joint. Overall, our study provides novel insights into critical determinants of inflammatory arthritis and may pave the way for dietary intervention trials in RA.

TAAR1 as an emerging target for the treatment of psychiatric disorders.

Trace amines, a group of amines expressed at the nanomolar level in the mammalian brain, can modulate monoamine transmission. The discovery of and the functional research on the trace amine-associated receptors (TAARs), especially the most well-characterized TAAR1, have largely facilitated our understanding of the function of the trace amine system in the brain. TAAR1 is expressed in the mammalian brain at a low level and widely distributed in the monoaminergic system, including the ventral tegmental area and substantial nigra, where the dopamine neurons reside in the mammalian brain. Growing in vitro and in vivo evidence has demonstrated that TAAR1 could negatively modulate monoamine transmission and play a crucial role in many psychiatric disorders, including schizophrenia, substance use disorders, sleep disorders, depression, and anxiety. Notably, in the last two decades, many studies have repeatedly confirmed the pharmacological effects of the selective TAAR1 ligands in various preclinical models of psychiatric disorders. Recent clinical trials of the dual TAAR1 and serotonin receptor agonist ulotaront also revealed a potential efficacy for treating schizophrenia. Here, we review the current understanding of the TAAR1 system and the recent advances in the elucidation of behavioral and physiological properties of TAAR1 agonists evaluated both in preclinical animal models and clinical trials. We also discuss the potential TAAR1-dependent signaling pathways and the cellular mechanisms underlying the inhibitory effects of TAAR1 activation on drug addiction. We conclude that TAAR1 is an emerging target for the treatment of psychiatric disorders.

Development of N-Terminally Modified Variants of the CXCR4-Antagonistic Peptide EPI-X4 for Enhanced Plasma Stability.

EPI-X4, a natural peptide CXCR4 antagonist, shows potential for treating inflammation and cancer, but its short plasma stability limits its clinical application. We aimed to improve the plasma stability of EPI-X4 analogues without compromising CXCR4 antagonism. Our findings revealed that only the peptide N-terminus is prone to degradation. Consequently, incorporating d-amino acids or acetyl groups in this region enhanced peptide stability in plasma. Notably, EPI-X4 leads 5, 27, and 28 not only retained their CXCR4 binding and antagonism but also remained stable in plasma for over 8 h. Molecular dynamic simulations showed that these modified analogues bind similarly to CXCR4 as the original peptide. To further increase their systemic half-lives, we conjugated these stabilized analogues with large polymers and albumin binders. These advances highlight the potential of the optimized EPI-X4 analogues as promising CXCR4-targeted therapeutics and set the stage for more detailed preclinical assessments.

Structural and signaling mechanisms of TAAR1 enabled preferential agonist design.

Trace amine-associated receptor 1 (TAAR1) senses a spectrum of endogenous amine-containing metabolites (EAMs) to mediate diverse psychological functions and is useful for schizophrenia treatment without the side effects of catalepsy. Here, we systematically profiled the signaling properties of TAAR1 activation and present nine structures of TAAR1-Gs/Gq in complex with EAMs, clinical drugs, and synthetic compounds. These structures not only revealed the primary amine recognition pocket (PARP) harboring the conserved acidic D3.32 for conserved amine recognition and "twin" toggle switch for receptor activation but also elucidated that targeting specific residues in the second binding pocket (SBP) allowed modulation of signaling preference. In addition to traditional drug-induced Gs signaling, Gq activation by EAM or synthetic compounds is beneficial to schizophrenia treatment. Our results provided a structural and signaling framework for molecular recognition by TAAR1, which afforded structural templates and signal clues for TAAR1-targeted candidate compounds design.

Trace Amine-Associated Receptors and Monoamine-Mediated Regulation of Insulin Secretion in Pancreatic Islets.

Currently, metabolic syndrome treatment includes predominantly pharmacological symptom relief and complex lifestyle changes. Trace amines and their receptor systems modulate signaling pathways of dopamine, norepinephrine, and serotonin, which are involved in the pathogenesis of this disorder. Trace amine-associated receptor 1 (TAAR1) is expressed in endocrine organs, and it was revealed that TAAR1 may regulate insulin secretion in pancreatic islet ß-cells. For instance, accumulating data demonstrate the positive effect of TAAR1 agonists on the dynamics of metabolic syndrome progression and MetS-associated disease development. The role of other TAARs (TAAR2, TAAR5, TAAR6, TAAR8, and TAAR9) in the islet's function is much less studied. In this review, we summarize the evidence of TAARs' contribution to the metabolic syndrome pathogenesis and regulation of insulin secretion in pancreatic islets. Additionally, by the analysis of public transcriptomic data, we demonstrate that TAAR1 and other TAAR receptors are expressed in the pancreatic islets. We also explore associations between the expression of TAARs mRNA and other genes in studied samples and demonstrate the deregulation of TAARs' functional associations in patients with metabolic diseases compared to healthy donors.

Ligand recognition and G-protein coupling of trace amine receptor TAAR1.

Trace-amine-associated receptors (TAARs), a group of biogenic amine receptors, have essential roles in neurological and metabolic homeostasis1. They recognize diverse endogenous trace amines and subsequently activate a range of G-protein-subtype signalling pathways2,3. Notably, TAAR1 has emerged as a promising therapeutic target for treating psychiatric disorders4,5. However, the molecular mechanisms underlying its ability to recognize different ligands remain largely unclear. Here we present nine cryo-electron microscopy structures, with eight showing human and mouse TAAR1 in a complex with an array of ligands, including the endogenous 3-iodothyronamine, two antipsychotic agents, the psychoactive drug amphetamine and two identified catecholamine agonists, and one showing 5-HT1AR in a complex with an antipsychotic agent. These structures reveal a rigid consensus binding motif in TAAR1 that binds to endogenous trace amine stimuli and two extended binding pockets that accommodate diverse chemotypes. Combined with mutational analysis, functional assays and molecular dynamic simulations, we elucidate the structural basis of drug polypharmacology and identify the species-specific differences between human and mouse TAAR1. Our study provides insights into the mechanism of ligand recognition and G-protein selectivity by TAAR1, which may help in the discovery of ligands or therapeutic strategies for neurological and metabolic disorders.

Evaluation of multienzyme supplementation and fiber levels on nutrient and energy digestibility of diets fed to gestating sows and growing pigs.

The objective was to investigate the effect of a multienzyme blend (MEblend) and inclusion level on apparent total tract digestibility (ATTD) of energy and nutrients, as well as ileal digestibility of crude protein (CP) and amino acids (AA) in gestation diets with low (LF) or high-dietary fiber (HF) fed to gestation sows. For comparison, growing pigs were fed the same HF diets to directly compare ATTD values with the gestating sows. In experiment 1, 45 gestating sows (parity 0 to 5; 187 ±â€…28 kg bodyweight; BW) were blocked by parity in a 2 × 3 factorial arrangement and fed 2.2 kg/d of the HF (17.5% neutral detergent fiber; NDF) or LF (13% NDF) diet and one of three levels of MEblend (0.0%, 0.08%, and 0.1%) to determine impacts of MEblend on ATTD. Twenty-seven growing pigs (initial 35.7 ±â€…3.32 kg BW) were fed the same HF diet (5% of BW) and one of three MEblend inclusions. The MEblend at both 0.08% and 0.1% increased ATTD of energy, NDF, and acid detergent fiber (ADF) (P < 0.05) in gestating sows but ATTD of total non-starch polysaccharides (NSP) and its residues were not affected. Sows fed HF, regardless of MEblend, had greater ATTD of NDF, xylose, and total NSP (P < 0.05) in comparison to grower pigs. In experiment 2, ileal cannulas were placed in 12 gestating sows (parity 0 to 2; BW 159 ±â€…12 kg) to determine apparent and standardized ileal digestibility (AID and SID) of AA and NSP. In a crossover design, sows were fed the same six diets, as in experiment 1, and a nitrogen-free diet during five periods of seven days each to achieve eight replicates per diet. There was no interaction between diet fiber level and MEblend inclusion. Supplementation of MEblend to gestating sow diets did not impact SID of CP and AA regardless of dietary fiber level. The SID of His, Ile, Lys, Phe, Thr, Trp, and Val were 3% to 6% lower (P < 0.09) in HF than LF independent of MEblend. Supplementation of MEblend did not impact AID of NSP components, but sows fed HF had higher AID of arabinose (LF: 26.5% vs. HF: 40.6%), xylose (LF: 3.5% vs. HF: 40.9%), and total NSP (LF: 25.9% vs. HF: 40.0%) compared to sows fed LF (P < 0.05). Dietary supplementation of MEblend increased ATTD of nutrients, NSP, and energy in diets fed to gestating sows regardless of inclusion level, with MEblend having a greater incremental increase in diets with lower NDF levels. Inclusion of MEblend impacted neither SID of AA nor AID of NSP in low- or high-fiber gestation diets, but high-fiber diet, negatively affected SID of AA.

Fiber-degrading enzymes have been extensively studied in growing pigs with minimal studies focusing on gestating sows; however, commercial gestating sow diets often contain more fiber than grower pig diets to stimulate the sensation of satiety without influencing weight gain. A challenge with dietary fiber is its hindrance on digestibility of nutrients. Supplementation of multienzyme blends increases nutrient digestibility of fibrous diets in grower pigs, but there is little data characterizing the effects of fiber-degrading enzymes in gestation diets for pregnant sows. In this study, inclusion of a multienzyme comprised of various carbohydrases and a protease in gestation diets increased apparent total tract digestibility of nutrients and energy for both gestating sows and growing pigs; however, digestibility of non-starch polysaccharides was only improved in growing pigs. Enzyme supplementation to gestating sow diets had limited impact on the ileal digestibility of nutrients, but ileal digestibility of amino acids and crude protein was reduced when gestating sows were fed diets higher in neutral detergent fiber. When formulating high-fiber diets for gestating sows and growing pigs using similar ingredients, it is critical to consider the differences in digestibility of fibrous components, particularly regarding ileal digestibility of amino acids.

Expression Pattern of Trace Amine-Associated Receptors during Differentiation of Human Pluripotent Stem Cells to Dopaminergic Neurons.

Trace amine-associated receptors (TAARs), which were discovered only in 2001, are known to be involved in the regulation of a spectrum of neuronal processes and may play a role in the pathogenesis of a number of neuropsychiatric diseases, such as schizophrenia and others. We have previously shown that TAARs also have interconnections with the regulation of neurogenesis and, in particular, with the neurogenesis of dopamine neurons, but the exact mechanisms of this are still unknown. In our work we analyzed the expression of TAARs (TAAR1, TAAR2, TAAR5, TAAR6, TAAR8 and TAAR9) in cells from the human substantia nigra and ventral tegmental areas and in human pluripotent stem cells at consecutive stages of their differentiation to dopaminergic neurons, using RNA sequencing data from open databases, and TaqMan PCR data from the differentiation of human induced pluripotent stem cells in vitro. Detectable levels of TAARs expression were found in cells at the pluripotent stages, and the dynamic of their expression had a trend of increasing with the differentiation and maturation of dopamine neurons. The expression of several TAAR types (particularly TAAR5) was also found in human dopaminergic neuron-enriched zones in the midbrain. This is the first evidence of TAARs expression during neuronal differentiation, which can help to approach an understanding of the role of TAARs in neurogenesis.