ResearchPad - amino-acid-analysis https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[Sequence-structure-function relationships in class I MHC: A local frustration perspective]]> https://www.researchpad.co/article/elastic_article_15751 Class I Major Histocompatibility Complex (MHC) binds short antigenic peptides with the help of Peptide Loading Complex (PLC), and presents them to T-cell Receptors (TCRs) of cytotoxic T-cells and Killer-cell Immunglobulin-like Receptors (KIRs) of Natural Killer (NK) cells. With more than 10000 alleles, human MHC (Human Leukocyte Antigen, HLA) is the most polymorphic protein in humans. This allelic diversity provides a wide coverage of peptide sequence space, yet does not affect the three-dimensional structure of the complex. Moreover, TCRs mostly interact with HLA in a common diagonal binding mode, and KIR-HLA interaction is allele-dependent. With the aim of establishing a framework for understanding the relationships between polymorphism (sequence), structure (conserved fold) and function (protein interactions) of the human MHC, we performed here a local frustration analysis on pMHC homology models covering 1436 HLA I alleles. An analysis of local frustration profiles indicated that (1) variations in MHC fold are unlikely due to minimally-frustrated and relatively conserved residues within the HLA peptide-binding groove, (2) high frustration patches on HLA helices are either involved in or near interaction sites of MHC with the TCR, KIR, or tapasin of the PLC, and (3) peptide ligands mainly stabilize the F-pocket of HLA binding groove.

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<![CDATA[Crystal structure of <i>Thermus thermophilus</i> methylenetetrahydrofolate dehydrogenase and determinants of thermostability]]> https://www.researchpad.co/article/elastic_article_13865 The elucidation of mechanisms behind the thermostability of proteins is extremely important both from the theoretical and applied perspective. Here we report the crystal structure of methylenetetrahydrofolate dehydrogenase (MTHFD) from Thermus thermophilus HB8, a thermophilic model organism. Molecular dynamics trajectory analysis of this protein at different temperatures (303 K, 333 K and 363 K) was compared with homologous proteins from the less temperature resistant organism Thermoplasma acidophilum and the mesophilic organism Acinetobacter baumannii using several data reduction techniques like principal component analysis (PCA), residue interaction network (RIN) analysis and rotamer analysis. These methods enabled the determination of important residues for the thermostability of this enzyme. The description of rotamer distributions by Gini coefficients and Kullback–Leibler (KL) divergence both revealed significant correlations with temperature. The emerging view seems to indicate that a static salt bridge/charged residue network plays a fundamental role in the temperature resistance of Thermus thermophilus MTHFD by enhancing both electrostatic interactions and entropic energy dispersion. Furthermore, this analysis uncovered a relationship between residue mutations and evolutionary pressure acting on thermophilic organisms and thus could be of use for the design of future thermostable enzymes.

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<![CDATA[Amino acids serve as an important energy source for adult flukes of <i>Clonorchis sinensis</i>]]> https://www.researchpad.co/article/elastic_article_13829 Clonorchiasis, closely related to cholangiocarcinoma and hepatocellular carcinoma, has led to a negative socioeconomic impact in global areas especially some Asian endemic regions. Owing to the emergence of drug resistance and hypersensitivity reactions after the massive and repeated use of praziquantel as well as the lack of effective vaccines, searching for new strategies that prevent and treat clonorchiasis has become an urgent matter. Clonorchis sinensis, the causative agent of clonorchiasis, long-term inhabits the microaerobic and limited-glucose environment of the bile ducts. Adequate nutrients are essential for adult flukes to resist the adverse condition and survive in the crowed habitat. Studies on energy metabolism of adult flukes are beneficial for further exploring host-parasite interactions and developing novel anti-parasitic drugs. Our results suggest that gluconeogenesis probably plays a vital role in energy metabolism of Clonorchis sinensis and exogenous amino acids might be an essential energy source for adult flukes to successfully survive in the host. Our foundational study opens a new avenue for explaining energy metabolism of Clonorchis sinensis and provides a valuable strategy that the gluconeogenesis pathway will be a potential and novel target for the prevention and treatment of clonorchiasis.

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<![CDATA[Physicochemical and biological evaluation of JR-131 as a biosimilar to a long-acting erythropoiesis-stimulating agent darbepoetin alfa]]> https://www.researchpad.co/article/Na789b0ff-1b14-409c-afa6-0c7a70fc7c42

Renal anemia is predominantly caused by a relative deficiency in erythropoietin (EPO). Conventional treatment for renal anemia includes the use of recombinant human EPO (rhEPO) or a long-acting erythropoiesis-activating agent named darbepoetin alfa, which is a modified rhEPO with a carbohydrate chain structure that differs from native hEPO. We have developed a biosimilar to darbepoetin alfa designated JR-131. Here, we comprehensively compare the physicochemical and biological characteristics of JR-131 to darbepoetin alfa. JR-131 demonstrated similar protein structure to the originator, darbepoetin alfa, by peptide mapping and circular dichroism spectroscopy. Additionally, mass spectroscopic analyses and capillary zone electrophoresis revealed similar glycosylation patterns between the two products. Human bone marrow-derived erythroblasts differentiated and proliferated to form colonies with JR-131 to a similar degree as darbepoetin alfa. Finally, JR-131 stimulated erythropoiesis and improved anemia in rats similarly to darbepoetin alfa. Our data show the similarity in physicochemical and biological properties of JR-131 to those of darbepoetin alfa, and JR-131 therefore represents a biosimilar for use in the treatment of renal anemia.

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<![CDATA[Sugar, amino acid and inorganic ion profiling of the honeydew from different hemipteran species feeding on Abies alba and Picea abies]]> https://www.researchpad.co/article/Neb889307-c28f-4dcd-8ba6-6ff5a5f28e34

Several hemipteran species feed on the phloem sap of plants and produce large amounts of honeydew that is collected by bees to produce honeydew honey. Therefore, it is important to know whether it is predominantly the hemipteran species or the host plant to influence the honeydew composition. This is particularly relevant for those botanical and zoological species from which the majority of honeydew honey originates. To investigate this issue, honeydew from two Cinara species located on Abies alba as well as from two Cinara and two Physokermes species located on Picea abies were collected. Phloem exudates of the host plants were also analyzed. Honeydew of all species contained different proportions of hexoses, sucrose, melezitose, erlose, and further di- and trisaccharides, whereas the phloem exudates of the host trees contained no trisaccharides. Moreover, the proportions of sugars differed significantly between hemipteran species feeding on the same tree species. Sucrose hydrolysis and oligosaccharide formation was shown in whole-body homogenates of aphids. The type of the produced oligosaccharides in the aphid-extracts correlated with the oligosaccharide composition in the honeydew of the different aphid species. The total contents of amino acids and inorganic ions in the honeydew were much lower than the sugar content. Glutamine and glutamate were predominant amino acids in the honeydew of all six hemipteran species and also in the phloem exudates of both tree species. Potassium was the dominant inorganic ion in all honeydew samples and also in the phloem exudate. Statistical analyses reveal that the sugar composition of honeydew is determined more by the hemipteran species than by the host plant. Consequently, it can be assumed that the sugar composition of honeydew honey is also more influenced by the hemipteran species than by the host tree.

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<![CDATA[Metabolomic profiling reveals correlations between spermiogram parameters and the metabolites present in human spermatozoa and seminal plasma]]> https://www.researchpad.co/article/5c76fe4fd5eed0c484e5b889

In 50% of all infertility cases, the male is subfertile or infertile, however, the underlying mechanisms are often unknown. Even when assisted reproductive procedures such as in vitro fertilization and intracytoplasmic sperm injection are performed, the causes of male factor infertility frequently remain elusive. Since the overall activity of cells is closely linked to their metabolic capacity, we analyzed a panel of 180 metabolites in human sperm and seminal plasma and elucidated their associations with spermiogram parameters. Therefore, metabolites from a group of 20 healthy donors were investigated using a targeted LC-MS/MS approach. The correlation analyses of the amino acids, biogenic amines, acylcarnitines, lysophosphatidylcholines, phosphatidylcholines, sphingomyelins and sugars from sperm and seminal plasma with standard spermiogram parameters revealed that metabolites in sperm are closely related to sperm motility, whereas those in seminal plasma are closely related to sperm concentration and morphology. This study provides essential insights into the metabolome of human sperm and seminal plasma and its associations with sperm functions. This metabolomics technique could be a promising screening tool to detect the factors of male infertility in cases where the cause of infertility is unclear.

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<![CDATA[Inherent versus induced protein flexibility: Comparisons within and between apo and holo structures]]> https://www.researchpad.co/article/5c5b52c9d5eed0c4842bd003

Understanding how ligand binding influences protein flexibility is important, especially in rational drug design. Protein flexibility upon ligand binding is analyzed herein using 305 proteins with 2369 crystal structures with ligands (holo) and 1679 without (apo). Each protein has at least two apo and two holo structures for analysis. The inherent variation in structures with and without ligands is first established as a baseline. This baseline is then compared to the change in conformation in going from the apo to holo states to probe induced flexibility. The inherent backbone flexibility across the apo structures is roughly the same as the variation across holo structures. The induced backbone flexibility across apo-holo pairs is larger than that of the apo or holo states, but the increase in RMSD is less than 0.5 Å. Analysis of χ1 angles revealed a distinctly different pattern with significant influences seen for ligand binding on side-chain conformations in the binding site. Within the apo and holo states themselves, the variation of the χ1 angles is the same. However, the data combining both apo and holo states show significant displacements. Upon ligand binding, χ1 angles are frequently pushed to new orientations outside the range seen in the apo states. Influences on binding-site variation could not be easily attributed to features such as ligand size or x-ray structure resolution. By combining these findings, we find that most binding site flexibility is compatible with the common practice in flexible docking, where backbones are kept rigid and side chains are allowed some degree of flexibility.

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<![CDATA[Pilot study of myocardial ischemia-induced metabolomic changes in emergency department patients undergoing stress testing]]> https://www.researchpad.co/article/5c5df30dd5eed0c484580c11

Background

The heart is a metabolically active organ, and plasma acylcarnitines are associated with long-term risk for myocardial infarction. We hypothesized that myocardial ischemia from cardiac stress testing will produce dynamic changes in acylcarnitine and amino acid levels compared to levels seen in matched control patients with normal stress tests.

Methods

We analyzed targeted metabolomic profiles in a pilot study of 20 case patients with inducible ischemia on stress testing from an existing prospectively collected repository of 357 consecutive patients presenting with symptoms of Acute Coronary Syndrome (ACS) in an Emergency Department (ED) observation unit between November 2012 and September 2014. We selected 20 controls matched on age, sex, and body-mass index (BMI). A peripheral blood sample was drawn <1 hour before stress testing and 2 hours after stress testing on each patient. We assayed 60 select acylcarnitines and amino acids by tandem mass spectrometry (MS/MS) using a Quattro Micro instrument (Waters Corporation, Milford, MA). Metabolite values were log transformed for skew. We then performed bivariable analysis for stress test outcome and both individual timepoint metabolite concentrations and stress-delta metabolite ratios (T2/T0). False discovery rates (FDR) were calculated for 60 metabolites while controlling for age, sex, and BMI. We built multivariable regularized linear models to predict stress test outcome from metabolomics data at times 0, 2 hours, and log ratio between these two. We used leave-one-out cross-validation to estimate the performance characteristics of the model.

Results

Nine of our 20 case subjects were male. Cases’ average age was 55.8, with an average BMI 29.5. Bivariable analysis identified 5 metabolites associated with positive stress tests (FDR < 0.2): alanine, C14:1-OH, C16:1, C18:2, C20:4. The multivariable regularized linear models built on T0 and T2 had Area Under the ROC Curve (AUC-ROC) between 0.5 and 0.55, however, the log(T2/T0) model yielded 0.625 AUC, with 65% sensitivity and 60% specificity. The top metabolites selected by the model were: Ala, Arg, C12-OH/C10-DC, C14:1-OH, C16:1, C18:2, C18:1, C20:4 and C18:1-DC.

Conclusions

Stress-delta metabolite analysis of patients undergoing stress testing is feasible. Future studies with a larger sample size are warranted.

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<![CDATA[Amino acid permeases in Cryptococcus neoformans are required for high temperature growth and virulence; and are regulated by Ras signaling]]> https://www.researchpad.co/article/5c57e680d5eed0c484ef3481

Cryptococcosis is an Invasive Fungal Infection (IFI) caused by Cryptococcus neoformans, mainly in immunocompromised patients. Therapeutic failure due to pathogen drug resistance, treatment inconstancy and few antifungal options is a problem. The study of amino acid biosynthesis and uptake represents an opportunity to explore possible development of novel antifungals. C. neoformans has 10 amino acids permeases, two of them (Aap3 and Aap7) not expressed at the conditions tested, and five were studied previously (Aap2, Aap4, Aap5, Mup1 and Mup3). Our previous results showed that Aap4 and Aap5 are major permeases with overlapping functions. The aap4Δ/aap5Δ double mutant fails to grow in amino acids as sole nitrogen source and is avirulent in animal model. Here, we deleted the remaining amino acid permeases (AAP1, AAP6, AAP8) that showed gene expression modulation by nutritional condition and created a double mutant (aap1Δ/aap2Δ). We studied the virulence attributes of these mutants and explored the regulatory mechanism behind amino acid uptake in C. neoformans. The aap1Δ/aap2Δ strain had reduced growth at 37°C in L-amino acids, reduced capsule production and was hypovirulent in the Galleria mellonella animal model. Our data, along with previous studies, (i) complement the analysis for all 10 amino acid permeases mutants, (ii) corroborate the idea that these transporters behave as global permeases, (iii) are required during heat and nutritional stress, and (iv) are important for virulence. Our study also indicates a new possible link between Ras1 signaling and amino acids uptake.

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<![CDATA[Software-aided workflow for predicting protease-specific cleavage sites using physicochemical properties of the natural and unnatural amino acids in peptide-based drug discovery]]> https://www.researchpad.co/article/5c3e50a9d5eed0c484d840db

Peptide drugs have been used in the treatment of multiple pathologies. During peptide discovery, it is crucially important to be able to map the potential sites of cleavages of the proteases. This knowledge is used to later chemically modify the peptide drug to adapt it for the therapeutic use, making peptide stable against individual proteases or in complex medias. In some other cases it needed to make it specifically unstable for some proteases, as peptides could be used as a system to target delivery drugs on specific tissues or cells. The information about proteases, their sites of cleavages and substrates are widely spread across publications and collected in databases such as MEROPS. Therefore, it is possible to develop models to improve the understanding of the potential peptide drug proteolysis. We propose a new workflow to derive protease specificity rules and predict the potential scissile bonds in peptides for individual proteases. WebMetabase stores the information from experimental or external sources in a chemically aware database where each peptide and site of cleavage is represented as a sequence of structural blocks connected by amide bonds and characterized by its physicochemical properties described by Volsurf descriptors. Thus, this methodology could be applied in the case of non-standard amino acid. A frequency analysis can be performed in WebMetabase to discover the most frequent cleavage sites. These results were used to train several models using logistic regression, support vector machine and ensemble tree classifiers to map cleavage sites for several human proteases from four different families (serine, cysteine, aspartic and matrix metalloproteases). Finally, we compared the predictive performance of the developed models with other available public tools PROSPERous and SitePrediction.

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<![CDATA[Seminal plasma amino acid profile in different breeds of chicken: Role of seminal plasma on sperm cryoresistance]]> https://www.researchpad.co/article/5c390b9ed5eed0c48491d873

Seminal plasma is a key biological fluid that modulates sperm function in the reproduction process. However, its role in sperm biotechnologies is scarce in poultry. The aims of the present study were to study the amino acids profile and total proteins of seminal plasma in 12 Spanish chicken breeds and to investigate the role of seminal plasma on cryoresistance of rooster sperm. To investigate the role of seminal plasma on cryoresistance, diluted pooled semen samples were cryopreserved in the presence and absence of seminal plasma. Glutamic acid was the most abundant free amino acid in seminal plasma, followed by alanine, serine, valine, and glycine. There was an influence of breed (P<0.05) on the percentage of viable sperm after freezing-thawing of samples with seminal plasma. Cluster analysis revealed that White Prat, Black Castellana, Blue Andaluza, Quail Castellana, and Red-Barred Vasca returned the best freezing-thawing response (good freezers). There was a positive correlation between seminal plasma concentrations of valine, isoleucine lysine, leucine and post thaw viability. The evaluation of fertilization capacity of frozen-thawed semen from the breeds White Prat (‘good freezer’) and Black-Red Andaluza (‘bad freezer’) showed that good freezer had higher fertility (20/68, 29.4%) compared to bad freezer breed (14/76, 18.4%), even if the difference was not significant (P = 0.08). The TUNEL assay revealed that freezing/thawing procedures in presence of seminal plasma provoked higher DNA fragmentation in most of the breeds, with a positive correlation between seminal alanine, valine, isoleucine, methionine, leucine, tyrosine, phenylalanine concentrations and DNA integrity. DNA fragmentation was lower in absence of seminal plasma and the breed effect on sperm viability was highly reduced. It is concluded that specific seminal plasma amino acids were associated with post-thaw percentage of viable sperm and DNA integrity. The removal of seminal plasma decreases the variability of the results and DNA fragmentation damages.

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<![CDATA[Amino acid residues in five separate HLA genes can explain most of the known associations between the MHC and primary biliary cholangitis]]> https://www.researchpad.co/article/5c0ed757d5eed0c484f13f5f

Primary Biliary Cholangitis (PBC) is a chronic autoimmune liver disease characterised by progressive destruction of intrahepatic bile ducts. The strongest genetic association is with HLA-DQA1*04:01, but at least three additional independent HLA haplotypes contribute to susceptibility. We used dense single nucleotide polymorphism (SNP) data in 2861 PBC cases and 8514 controls to impute classical HLA alleles and amino acid polymorphisms using state-of-the-art methodologies. We then demonstrated through stepwise regression that association in the HLA region can be largely explained by variation at five separate amino acid positions. Three-dimensional modelling of protein structures and calculation of electrostatic potentials for the implicated HLA alleles/amino acid substitutions demonstrated a correlation between the electrostatic potential of pocket P6 in HLA-DP molecules and the HLA-DPB1 alleles/amino acid substitutions conferring PBC susceptibility/protection, highlighting potential new avenues for future functional investigation.

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<![CDATA[Characteristics of interactions at protein segments without non-local intramolecular contacts in the Protein Data Bank]]> https://www.researchpad.co/article/5c1966f9d5eed0c484b537eb

The principle of three-dimensional protein structure formation is a long-standing conundrum in structural biology. A globular domain of a soluble protein is formed by a network of atomic contacts among amino acid residues, but regions without intramolecular non-local contacts are often observed in the protein structure, especially in loop, linker, and peripheral segments with secondary structures. Although these regions can play key roles for protein function as interfaces for intermolecular interactions, their nature remains unclear. Here, we termed protein segments without non-local contacts as floating segments and sought them in tens of thousands of entries in the Protein Data Bank. As a result, we found that 0.72% of residues are in floating segments. Regarding secondary structural elements, coil structures are enriched in floating segments, especially for long segments. Interactions with polypeptides and polynucleotides, but not chemical compounds, are enriched in floating segments. The amino acid preferences of floating segments are similar to those of surface residues, with exceptions; the small side chain amino acids, Gly and Ala, are preferred, and some charged side chains, Arg and His, are disfavored for floating segments compared to surface residues. Our comprehensive characterization of floating segments may provide insights into understanding protein sequence-structure-function relationships.

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<![CDATA[Extended cleavage specificities of mast cell proteases 1 and 2 from golden hamster: Classical chymase and an elastolytic protease comparable to rat and mouse MCP-5]]> https://www.researchpad.co/article/5c12cf42d5eed0c484914256

Serine proteases constitute the major protein content of mast cell secretory granules. Here we present the extended cleavage specificity of two such proteases from the golden hamster, Mesocricetus auratus. Analysis by phage display technique showed that one of them (HAM1) is a classical chymase with a specificity similar to the human mast cell chymase. However, in contrast to the human chymase, it does not seem to have a particular preference for any of the three aromatic amino acids, Phe, Tyr and Trp, in the P1 position of substrates. HAM1 also efficiently cleaved after Leu similarly to human and many other mast cell chymases. We observed only a 3-fold lower cleavage activity on Leu compared to substrates with P1 aromatic amino acids. Chymotryptic enzymes seem to be characteristic for connective tissue mast cells in mammalian species from opossums to humans, which indicates a very central role of these enzymes in mast cell biology. HAM1 also seems to have the strongest preference for negatively charged amino acids in the P2´position of all mast cell chymases so far characterized. The second hamster chymase, HAM2, is an elastolytic in its activity, similarly to the α-chymases in rats and mice (rMCP-5 and mMCP-5, respectively). The presence of an α-chymase that developed elastase activity thereby seems to be a relatively early modification of the α-chymase within the rodent branch of the mammalian evolutionary tree.

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<![CDATA[Base composition is the primary factor responsible for the variation of amino acid usage in zebra finch (Taeniopygia guttata)]]> https://www.researchpad.co/article/5c117b88d5eed0c4846998fc

In the present study, we carried out an examination of the amino acid usage in the zebra finch (Taeniopygia guttata) proteome. We found that tRNA abundance, base composition, hydrophobicity and aromaticity, protein second structure, cysteine residue (Cys) content and protein molecular weight had significant impact on the amino acid usage of the zebra finch. The above factors explained the total variability of 22.85%, 25.37%, 10.91%, 5.06%, 4.21%, and 3.14%, respectively. Altogether, approximately 70% of the total variability in zebra finch could be explained by such factors. Comparison of the amino acid usage between zebra finch, chicken (Gallus gallus) and human (Homo sapiens) suggested that the average frequency of various amino acid usage is generally consistent among them. Correspondence analysis indicated that base composition was the primary factor affecting the amino acid usage in zebra finch. This trend was different from chicken, but similar to human. Other factors affecting the amino acid usage in zebra finch, such as isochore structure, protein second structure, Cys frequency and protein molecular weight also showed the similar trends with human. We do not know whether the similar amino acid usage trend between human and zebra finch is related to the distinctive neural and behavioral traits, but it is worth studying in depth.

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<![CDATA[Metabolomics approach in the investigation of depression biomarkers in pharmacologically induced immune-related depression]]> https://www.researchpad.co/article/5c0993c5d5eed0c4842ad8ac

Background

The aim of this study was to identify previously unrecognised biological pathways and biomarkers that might expand the inflammatory hypothesis of depression.

Methods

Broad metabolomics analyses in plasma samples from 31 chronic hepatitis C-infected patients with and without immune-related depression were carried out using the Absolute IDQ p180 kit—a targeted metabolomics approach of combined direct flow injection and liquid chromatography that measures acylcarnitines, amino acids, biogenic amines, glycerophospholipids, sphingolipids, and sugars.

Results

The measurements showed that the average concentration of the branched-chain amino acid isoleucine was significantly lower in depressive HCV patients in comparison to non-depressive HCV patients [depression group: Median 51.35 (43.4–60.2 μmol/L) vs. Median 62.10 (38.4–81.7 μmol/L); U = -2.958; p = 0.002]. All other amino acids, acylcarnitines, biogenic amines, glycerophospholipids, sphingolipids, sugars, liver enzymes and thyroid levels showed no statistically significant differences.

Conclusions

The results of the present study suggest that the branched-chain amino acid isoleucine might play a role in the pathophysiology of immune-related major depression, which expands existing knowledge about inflammatory hypothesis of depression.

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<![CDATA[The influence of feeding behaviour on growth performance, carcass and meat characteristics of growing pigs]]> https://www.researchpad.co/article/5bce3d1540307c69b197f500

This study investigated the effect of the feeding behaviour on growth performance, and carcass and meat characteristics of 96 barrows fed ad libitum or restrictively with high or low amino acids (AA) diets according to a 2 × 2 factorial design. The feeding behaviour traits were measured with automated feeders. From 86 kg BW, half of the pigs were given feeds with high indispensable (AA) contents, while the other half received feeds with indispensable AA contents reduced by 9% in early finishing (86–118 kg BW) and by 18% in late finishing (118–145 kg BW). Body lipid and protein retentions were estimated from BW and backfat depth measures recorded at the beginning and end of each period. Pigs were slaughtered at 145 kg BW and carcass and meat quality data were recorded. Phenotypic correlations among feeding behaviours, growth performances, and carcass and meat traits were computed from all the data after adjustment for the effects of feeding treatments. As feeding rate was the behavioural trait most highly correlated with performance and carcass traits, the records of each pig were classified into feeding rate tertiles. Then, the data were statistically analysed using a mixed model, which included feed restriction (FR), AA reduction (AAR), the FR × AAR interaction and the feeding rate tertile as fixed factors, and pen as a random factor. Pigs eating faster (52.1 to 118.9 g/min) had significantly greater final body weights (16%), average daily weight gains (27%), estimated protein gains (22%), estimated lipid retention (46%), carcass weights (16%), weights of lean cuts (14%), weights of fat cuts (21%), proportions of fat in the carcass (14%), and 4% lower proportions of carcass lean cuts than pigs eating slowly (12.6 to 38.2 g/min). Manipulating the eating rate, through management or genetic strategies, could affect feed intake and subsequent growth performance, hence carcass quality, but have little influence on feed efficiency.

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<![CDATA[Changes in the free amino acid composition of Capsicum annuum (pepper) leaves in response to Myzus persicae (green peach aphid) infestation. A comparison with water stress]]> https://www.researchpad.co/article/5c032defd5eed0c4844f899d

Amino acids play a central role in aphid-plant interactions. They are essential components of plant primary metabolism, function as precursors for the synthesis of defense-related specialized metabolites, and are major growth-limiting nutrients for aphids. To quantify changes in the free amino acid content of pepper (Capsicum annuum L.) leaves in response to green peach aphid (Myzus persicae Sulzer) feeding, plants were infested with a low (20 aphids/plant) or a high (200 aphids/plant) aphid density in time-course experiments ranging from 3 hours to 7 days. A parallel experiment was conducted with pepper plants that had been subjected to water stress. Factor Analysis of Mixed Data revealed a significant interaction of time x density in the free amino acid response of aphid-infested leaves. At low aphid density, M. persicae did not trigger a strong response in pepper leaves. Conversely, at high density, a large increase in total free amino acid content was observed and specific amino acids peaked at different times post-infestation. Comparing aphid-infested with water-stressed plants, most of the observed differences were quantitative. In particular, proline and hydroxyproline accumulated dramatically in response to water stress, but not in response to aphid infestation. Some additional differences and commonalities between the two stress treatments are discussed.

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<![CDATA[In Silico Analysis of the Structural and Biochemical Features of the NMD Factor UPF1 in Ustilago maydis]]> https://www.researchpad.co/article/5989daedab0ee8fa60bbfc25

The molecular mechanisms regulating the accuracy of gene expression are still not fully understood. Among these mechanisms, Nonsense-mediated Decay (NMD) is a quality control process that detects post-transcriptionally abnormal transcripts and leads them to degradation. The UPF1 protein lays at the heart of NMD as shown by several structural and functional features reported for this factor mainly for Homo sapiens and Saccharomyces cerevisiae. This process is highly conserved in eukaryotes but functional diversity can be observed in various species. Ustilago maydis is a basidiomycete and the best-known smut, which has become a model to study molecular and cellular eukaryotic mechanisms. In this study, we performed in silico analysis to investigate the structural and biochemical properties of the putative UPF1 homolog in Ustilago maydis. The putative homolog for UPF1 was recognized in the annotated genome for the basidiomycete, exhibiting 66% identity with its human counterpart at the protein level. The known structural and functional domains characteristic of UPF1 homologs were also found. Based on the crystal structures available for UPF1, we constructed different three-dimensional models for umUPF1 in order to analyze the secondary and tertiary structural features of this factor. Using these models, we studied the spatial arrangement of umUPF1 and its capability to interact with UPF2. Moreover, we identified the critical amino acids that mediate the interaction of umUPF1 with UPF2, ATP, RNA and with UPF1 itself. Mutating these amino acids in silico showed an important effect over the native structure. Finally, we performed molecular dynamic simulations for UPF1 proteins from H. sapiens and U. maydis and the results obtained show a similar behavior and physicochemical properties for the protein in both organisms. Overall, our results indicate that the putative UPF1 identified in U. maydis shows a very similar sequence, structural organization, mechanical stability, physicochemical properties and spatial organization in comparison to the NMD factor depicted for Homo sapiens. These observations strongly support the notion that human and fungal UPF1 could perform equivalent biological activities.

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<![CDATA[Unifying view of mechanical and functional hotspots across class A GPCRs]]> https://www.researchpad.co/article/5989db54ab0ee8fa60bdd0a8

G protein-coupled receptors (GPCRs) are the largest superfamily of signaling proteins. Their activation process is accompanied by conformational changes that have not yet been fully uncovered. Here, we carry out a novel comparative analysis of internal structural fluctuations across a variety of receptors from class A GPCRs, which currently has the richest structural coverage. We infer the local mechanical couplings underpinning the receptors’ functional dynamics and finally identify those amino acids whose virtual deletion causes a significant softening of the mechanical network. The relevance of these amino acids is demonstrated by their overlap with those known to be crucial for GPCR function, based on static structural criteria. The differences with the latter set allow us to identify those sites whose functional role is more clearly detected by considering dynamical and mechanical properties. Of these sites with a genuine mechanical/dynamical character, the top ranking is amino acid 7x52, a previously unexplored, and experimentally verifiable key site for GPCR conformational response to ligand binding.

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