Ribonucleotides differentially modulate oral glutamate detection thresholds.

The savory or umami taste of the amino acid glutamate is synergistically enhanced by the addition of the purines inosine 5'-monophosphate (IMP) and guanosine 5'-monophosphate (GMP) disodium salt. We hypothesized that the addition of purinergic ribonucleotides, along with the pyrimidine ribonucleotides, would decrease the absolute detection threshold of (increase sensitivity to) l-glutamic acid potassium salt (MPG). To test this, we measured both the absolute detection threshold of MPG alone and with a background level (3 mM) of 5 different 5'-ribonucleotides. The addition of the 3 purines IMP, GMP, and adenosine 5'-monophosphate (AMP) lowered the MPG threshold in all participants (P < 0.001), indicating they are positive modulators or enhancers of glutamate taste. The average detection threshold of MPG was 2.08 mM, and with the addition of IMP, the threshold was decreased by approximately 1.5 orders of magnitude to 0.046 mM. In contrast to the purines, the pyrimidines uridine 5'-monophosphate (UMP) and cytidine 5'-monophosphate (CMP) yielded different results. CMP reliably raised glutamate thresholds in 10 of 17 subjects, suggesting it is a negative modulator or diminisher of glutamate taste for them. The rank order of effects on increasing sensitivity to glutamate was IMP > GMP> AMP >> UMP// CMP. These data confirm that ribonucleotides are modulators of glutamate taste, with purines enhancing sensitivity and pyrimidines displaying variable and even negative modulatory effects. Our ability to detect the co-occurrence of glutamate and purines is meaningful as both are relatively high in evolutionarily important sources of nutrition, such as insects and fermented foods.

Metabolomics reveals inosine 5'-monophosphate is increased during mice adipocyte browning.

Adipocyte browning is one of the potential strategies for the prevention of obesity-related metabolic syndromes, but it is a complex process. Although previous studies make it increasingly clear that several transcription factors and enzymes are essential to induce browning, it is unclear what dynamic and metabolic changes occur in induction of browning. Here, we analyzed the effect of a beta-adrenergic receptor agonist (CL316243, accelerator of browning) on metabolic change in mice adipose tissue and plasma using metabolome analysis and speculated that browning is regulated partly by inosine 5'-monophosphate (IMP) metabolism. To test this hypothesis, we investigated whether Ucp-1, a functional marker of browning, mRNA expression is influenced by IMP metabolism using immortalized adipocytes. Our study showed that mycophenolic acid, an IMP dehydrogenase inhibitor, increases the mRNA expression of Ucp-1 in immortalized adipocytes. Furthermore, we performed a single administration of mycophenolate mofetil, a prodrug of mycophenolic acid, to mice and demonstrated that mycophenolate mofetil induces adipocyte browning and miniaturization of adipocyte size, leading to adipose tissue weight loss. These findings showed that IMP metabolism has a significant effect on adipocyte browning, suggesting that the regulator of IMP metabolism has the potential to prevent obesity.

AICAR transformylase/IMP cyclohydrolase (ATIC) is essential for de novo purine biosynthesis and infection by Cryptococcus neoformans.

The fungal pathogen Cryptococcus neoformans is a leading cause of meningoencephalitis in the immunocompromised. As current antifungal treatments are toxic to the host, costly, limited in their efficacy, and associated with drug resistance, there is an urgent need to identify vulnerabilities in fungal physiology to accelerate antifungal discovery efforts. Rational drug design was pioneered in de novo purine biosynthesis as the end products of the pathway, ATP and GTP, are essential for replication, transcription, and energy metabolism, and the same rationale applies when considering the pathway as an antifungal target. Here, we describe the identification and characterization of C. neoformans 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) transformylase/5'-inosine monophosphate cyclohydrolase (ATIC), a bifunctional enzyme that catalyzes the final two enzymatic steps in the formation of the first purine base inosine monophosphate. We demonstrate that mutants lacking the ATIC-encoding ADE16 gene are adenine and histidine auxotrophs that are unable to establish an infection in a murine model of virulence. In addition, our assays employing recombinantly expressed and purified C. neoformans ATIC enzyme revealed Km values for its substrates AICAR and 5-formyl-AICAR are 8-fold and 20-fold higher, respectively, than in the human ortholog. Subsequently, we performed crystallographic studies that enabled the determination of the first fungal ATIC protein structure, revealing a key serine-to-tyrosine substitution in the active site, which has the potential to assist the design of fungus-specific inhibitors. Overall, our results validate ATIC as a promising antifungal drug target.

Kinetic Characterization and Inhibition of Trypanosoma cruzi Hypoxanthine-Guanine Phosphoribosyltransferases.

Chagas disease, caused by the parasitic protozoan Trypanosoma cruzi, affects over 8 million people worldwide. Current antiparasitic treatments for Chagas disease are ineffective in treating advanced, chronic stages of the disease, and are noted for their toxicity. Like most parasitic protozoa, T. cruzi is unable to synthesize purines de novo, and relies on the salvage of preformed purines from the host. Hypoxanthine-guanine phosphoribosyltransferases (HGPRTs) are enzymes that are critical for the salvage of preformed purines, catalyzing the formation of inosine monophosphate (IMP) and guanosine monophosphate (GMP) from the nucleobases hypoxanthine and guanine, respectively. Due to the central role of HGPRTs in purine salvage, these enzymes are promising targets for the development of new treatment methods for Chagas disease. In this study, we characterized two gene products in the T. cruzi CL Brener strain that encodes enzymes with functionally identical HGPRT activities in vitro: TcA (TcCLB.509693.70) and TcC (TcCLB.506457.30). The TcC isozyme was kinetically characterized to reveal mechanistic details on catalysis, including identification of the rate-limiting step(s) of catalysis. Furthermore, we identified and characterized inhibitors of T. cruzi HGPRTs originally developed as transition-state analogue inhibitors (TSAIs) of Plasmodium falciparum hypoxanthine-guanine-xanthine phosphoribosyltransferase (PfHGXPRT), where the most potent compound bound to T. cruzi HGPRT with low nanomolar affinity. Our results validated the repurposing of TSAIs to serve as selective inhibitors for orthologous molecular targets, where primary and secondary structures as well as putatively common chemical mechanisms are conserved.

Disorders of purine biosynthesis metabolism.

Purines are essential molecules that are components of vital biomolecules, such as nucleic acids, coenzymes, signaling molecules, as well as energy transfer molecules. The de novo biosynthesis pathway starts from phosphoribosylpyrophosphate (PRPP) and eventually leads to the synthesis of inosine monophosphate (IMP) by means of 10 sequential steps catalyzed by six different enzymes, three of which are bi-or tri-functional in nature. IMP is then converted into guanosine monophosphate (GMP) or adenosine monophosphate (AMP), which are further phosphorylated into nucleoside di- or tri-phosphates, such as GDP, GTP, ADP and ATP. This review provides an overview of inborn errors of metabolism pertaining to purine synthesis in humans, including either phosphoribosylpyrophosphate synthetase (PRS) overactivity or deficiency, as well as adenylosuccinate lyase (ADSL), 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase (ATIC), phosphoribosylaminoimidazole succinocarboxamide synthetase (PAICS), and adenylosuccinate synthetase (ADSS) deficiencies. ITPase deficiency is being described as well. The clinical spectrum of these disorders is broad, including neurological impairment, such as psychomotor retardation, epilepsy, hypotonia, or microcephaly; sensory involvement, such as deafness and visual disturbances; multiple malformations, as well as muscle presentations or consequences of hyperuricemia, such as gouty arthritis or kidney stones. Clinical signs are often nonspecific and, thus, overlooked. It is to be hoped that this is likely to be gradually overcome by using sensitive biochemical investigations and next-generation sequencing technologies.

Supplementation effects of a kokumi substance, γ-Glu-Val-Gly, on the ingestion of basic taste solutions in rats.

In addition to the well-accepted taste receptors corresponding to the 6 basic taste qualities, sweet, salty, sour, bitter, umami, and fatty, another type of taste receptor, calcium-sensing receptor (CaSR), is located in taste bud cells. CaSR is called the kokumi receptor because its agonists induce koku (or kokumi), a Japanese word meaning the enhancement of flavor characteristics, such as thickness, mouthfulness, and continuity. Kokumi is an important factor in enhancing food palatability. γ-Glu-Val-Gly (EVG) is the most potent agonist of CaSR, which induces a strong kokumi flavor. However, no behavioral studies have been documented in animals using EVG. Here, we show that EVG at low concentrations that do not elicit a taste of its own enhances preferences for umami, fat, and sweet taste solutions in rats. An increased preference for inosine monophosphate (IMP) and Intralipos was the most dominant effect. NPS-2143, an antagonist of CaSR, abolished the additive effect of EVG on IMP and Intralipos solutions. These effects of EVG on taste stimuli are thought to occur in the oral cavity, because the effects of EVG were confirmed in a brief exposure test. The additive effects on IMP and Intralipos remained after the transection of the chorda tympani, indicating that these effects also occur in the palate and/or posterior part of the tongue. Moreover, the additive effects of EVG were verified in electrophysiological taste nerve responses. These results may partially provide the underlying mechanisms for EVG to induce kokumi flavor in humans.

Inosine monophosphate and inosine differentially regulate endotoxemia and bacterial sepsis.

Inosine monophosphate (IMP) is the intracellular precursor for both adenosine monophosphate and guanosine monophosphate and thus plays a central role in intracellular purine metabolism. IMP can also serve as an extracellular signaling molecule, and can regulate diverse processes such as taste sensation, neutrophil function, and ischemia-reperfusion injury. How IMP regulates inflammation induced by bacterial products or bacteria is unknown. In this study, we demonstrate that IMP suppressed tumor necrosis factor (TNF)-α production and augmented IL-10 production in endotoxemic mice. IMP exerted its effects through metabolism to inosine, as IMP only suppressed TNF-α following its CD73-mediated degradation to inosine in lipopolysaccharide-activated macrophages. Studies with gene targeted mice and pharmacological antagonism indicated that A2A , A2B, and A3 adenosine receptors are not required for the inosine suppression of TNF-α production. The inosine suppression of TNF-α production did not require its metabolism to hypoxanthine through purine nucleoside phosphorylase or its uptake into cells through concentrative nucleoside transporters indicating a role for alternative metabolic/uptake pathways. Inosine augmented IL-ß production by macrophages in which inflammasome was activated by lipopolysaccharide and ATP. In contrast to its effects in endotoxemia, IMP failed to affect the inflammatory response to abdominal sepsis and pneumonia. We conclude that extracellular IMP and inosine differentially regulate the inflammatory response.

Research progress on inosine monophosphate deposition mechanism in chicken muscle.

With the continuous improvements in human diet, there is an ever-increasing demand for high-quality chicken, so it is particularly important for poultry breeders to carry out the breeding of high-quality broilers in a timely fashion. Inosine monophosphate (IMP) is a flavor-enhancing substance, which plays a critical role in the umami taste of the muscle, making the content of IMP an important umami taste indicator. Currently, research on the deposition mechanism of IMP in chicken is not only necessary for chicken breeders to promote the production of high-quality meat and poultry but also to meet the human demand for chicken meat. In this paper, the research history of IMP, its structure and taste mechanisms, the pathway and influencing factors of de novo IMP synthesis, and the key genes regulating IMP synthesis and metabolism are briefly summarized. Our aim was to lay a theoretical foundation and provide scientific background and research directions for further research on high-quality broiler breeding.

Inosine monophosphate dehydrogenase activity and mycophenolate pharmacokinetics in children with nephrotic syndrome treated with mycophenolate mofetil.

Some studies have shown that the area under the concentration-time curve (AUC) of mycophenolic acid (MPA) should be higher for children with nephrotic syndrome (NS) than after renal transplantation. The pharmacodynamic aspect of MPA, the activity of inosine monophosphate dehydrogenase (IMPDH), has not been studied in children with NS. The study included 21 children (4-16 years old) with NS treated with mycophenolate mofetil. MPA and its glucuronide plasma concentrations were determined using validated high-performance liquid chromatography-ultraviolet (HPLC-UV). The separate HPLC-UV method was applied for IMPDH activity determination. The variability was expressed by the coefficient of variation (CV). IMPDH activity and MPA concentration (Ctrough ) before the morning dose amounted to 29.95 µmol s-1  mol-1 adenosine monophosphate (AMP) (range, 6.71-98.60 µmol s-1  mol-1 AMP) and 1.72 µg/mL (range, 0.39-4.34 µg/mL), respectively, whereas the area under the effect-time curve from 0 to 4 h and MPA AUC0-4 were 130.36 µmol s-1  mol-1 AMP × h (range, 23.58-306.57 µmol s-1  mol-1 AMP × h) and 24.63 µg h/mL (range, 12.21-67.48 µg h/mL), respectively. IMPDH activity decreased concomitantly with MPA concentration increase, however, the variability of the pharmacodynamic parameters was greater than of the pharmacokinetics. The median degree of maximum IMPDH inhibition was 61%. MPA Ctrough and predicted AUC were lower than in our previous study. Only a few MPA pharmacokinetic parameters correlated with the pharmacodynamics. IMPDH activity did not correlate with the children's age and did not differ between boys and girls. MPA clearance was the highest in younger children (median, 10.54 L/m2 /h) and cholesterol correlated negatively with the children's age (r = -0.659, P = 0.003). IMPDH minimum activity and the degree of maximum IMPDH inhibition were similar to those obtained in renal transplant recipients. IMPDH activity does not undergo developmental or gender-specific regulation in children with NS. MPA underexposure might be more frequent in younger children, especially with high cholesterol and triglycerides levels because of high MPA clearance.

The Application of In Silico Methods on Umami Taste Receptor.

The umami taste receptor is a heterodimer composed of two members of the T1R taste receptor family: T1R1 (taste receptor type 1 member 1) and T1R3 (taste receptor type 1 member 3). Taste receptor T1R1-T1R3 can be activated, or modulated, by binding to several natural ligands, such as L-glutamate, inosine-5'-monophosphate (IMP), and guanosine-5'-monophosphate (GMP). Because no structure of the umami taste receptor has been solved until now, in silico techniques, such as homology modelling, molecular docking, and molecular dynamics (MD) simulations, are used to generate a 3D structure model of this receptor and to understand its molecular mechanisms. The purpose of this chapter is to highlight how computational methods can provide a better deciphering of the mechanisms of action of umami ligands in activating the umami taste receptors leading to advancements in the taste research field.

miRNA-mRNA associations with inosine monophosphate specific deposition in the muscle of Jingyuan chicken.

1. Inosine monophosphate (IMP), is an essential component for meat flavour and microRNAs (miRNAs) play a vital role in its post-transcriptional regulation. However, the mechanism of how miRNA expression affects muscle-specific IMP deposition is unclear.2. The following study performed transcriptome sequencing and bioinformatics analysis of breast and leg muscle, which have significantly different IMP content in Jingyuan chicken. The differential miRNA-mRNAs were screened out and correlation analysis with IMP content was performed.3. A total of 39 differentially expressed miRNAs (DE miRNAs) and 666 differentially expressed mRNAs (DE mRNAs) were identified between breast muscles and leg muscles. Using miRNA-mRNA integrated analysis, 29 miRNA-target gene pairs were obtained, composed of 13 DE miRNAs and 28 DE mRNAs. Next, purine metabolism, glycolysis/gluconeogenesis, pyruvate metabolism and the biosynthesis of amino acid pathways as necessary for muscle IMP-specific deposition were identified. The differentially expressed gene PKM2, which was significantly enriched in all four pathways, is involved in IMP anabolism in the form of energy metabolism and enzyme activity regulation. The correlation analysis suggested that the gga-miR-107-3p-KLHDC2 negative interaction may be a key regulator in IMP deposition.4. This study explores the functional mechanism of IMP-specific deposition in Jingyuan chicken muscles at the miRNA and mRNA levels and highlights multiple candidate miRNAs and mRNAs for molecular-assisted breeding.

Reprogramming of macrophages employing gene regulatory and metabolic network models.

Upon exposure to different stimuli, resting macrophages undergo classical or alternative polarization into distinct phenotypes that can cause fatal dysfunction in a large range of diseases, such as systemic infection leading to sepsis or the generation of an immunosuppressive tumor microenvironment. Investigating gene regulatory and metabolic networks, we observed two metabolic switches during polarization. Most prominently, anaerobic glycolysis was utilized by M1-polarized macrophages, while the biosynthesis of inosine monophosphate was upregulated in M2-polarized macrophages. Moreover, we observed a switch in the urea cycle. Gene regulatory network models revealed E2F1, MYC, PPARγ and STAT6 to be the major players in the distinct signatures of these polarization events. Employing functional assays targeting these regulators, we observed the repolarization of M2-like cells into M1-like cells, as evidenced by their specific gene expression signatures and cytokine secretion profiles. The predicted regulators are essential to maintaining the M2-like phenotype and function and thus represent potential targets for the therapeutic reprogramming of immunosuppressive M2-like macrophages.

Characterizing perceptions of an abbreviated ISMP Medication Safety Self Assessment for community pharmacies.

OBJECTIVES: This study sought to characterize users' perceptions of, and identify the average time needed to complete a newly abbreviated version of the Institute for Safe Medication Practices Medication Safety Self Assessment for Community and Ambulatory Pharmacy (MSSA-CAP). METHODS: This study took place within a large, national, nonprofit, faith-based health system. An abbreviated version of ISMP's MSSA-CAP was developed through an iterative process by researchers and the health system's medication safety officers (MSOs, i.e., the assessment tool's 'users'). Retained items included those with nonoverlapping (1) bolded or keywords; (2) "quick asks," answerable without external information; or (3) were relevant to community pharmacy practice (i.e., nonambulatory care-specific). During site visits, MSOs assessed the organization's community pharmacies with the abbreviated tool. Users completed pre- and post-visit surveys asking their perceptions of and time spent using the full and abbreviated versions of the MSSA-CAP. Results were analyzed using descriptive statistics. RESULTS: Sixty of the original MSSA-CAP's 216 items were retained. Between August to December 2019, six MSO users assessed 59 community pharmacies across 10 states with the abbreviated assessment tool. On the average, users reported needing 86.1 ± 35.4 minutes to complete the abbreviated assessment. Sixty-seven percent of users agreed or strongly agreed that the abbreviated assessment was of a good length, compared with only 17% for the original full assessment. Collectively, assessed community pharmacies scored highest on MSSA-CAP items related to Physical Environment and Prescription Labels, and lowest on Hard Stops and Proactive Risk Assessments. CONCLUSION: Streamlining items in medication safety assessment tools may be useful in overcoming time barriers to implementation in community pharmacies.

Small Molecule CD38 Inhibitors: Synthesis of 8-Amino-N1-inosine 5'-monophosphate, Analogues and Early Structure-Activity Relationship.

Although a monoclonal antibody targeting the multifunctional ectoenzyme CD38 is an FDA-approved drug, few small molecule inhibitors exist for this enzyme that catalyzes inter alia the formation and metabolism of the N1-ribosylated, Ca2+-mobilizing, second messenger cyclic adenosine 5'-diphosphoribose (cADPR). N1-Inosine 5'-monophosphate (N1-IMP) is a fragment directly related to cADPR. 8-Substituted-N1-IMP derivatives, prepared by degradation of cyclic parent compounds, inhibit CD38-mediated cADPR hydrolysis more efficiently than related cyclic analogues, making them attractive for inhibitor development. We report a total synthesis of the N1-IMP scaffold from adenine and a small initial compound series that facilitated early delineation of structure-activity parameters, with analogues evaluated for inhibition of CD38-mediated hydrolysis of cADPR. The 5'-phosphate group proved essential for useful activity, but substitution of this group by a sulfonamide bioisostere was not fruitful. 8-NH2-N1-IMP is the most potent inhibitor (IC50 = 7.6 µM) and importantly HPLC studies showed this ligand to be cleaved at high CD38 concentrations, confirming its access to the CD38 catalytic machinery and demonstrating the potential of our fragment approach.

The potential effects of anticonvulsant drugs on neuropeptides and neurotrophins in pentylenetetrazol kindled seizures in the rat.

Purpose: Neuropeptides and neurotrophic factors are thought to be involved in epileptogenesis. This study aims to investigate the potential effects of anticonvulsant drugs on neuropeptides (galanin and neuropeptide Y) and neurotrophic factors (BDNF and NGF) in pentylenetetrazol (PTZ)-kindled seizures in the rat.Methods: Forty-eight adult male Sprague-Dawley rats were included in the study. The animals were divided into 8 groups of six rats. Group 1 was defined as naïve control, and received no medication. Group 2 (PTZ + saline) was treated with sub-convulsive doses of PTZ (35 mg/kg) and saline i.p. for 14 days. For anticonvulsant treatments, Groups 3-8 were treated with 200 mg/kg levetiracetam (PTZ + LEV), 1 mg/kg midazolam (PTZ + MDZ), 80 mg/kg phenytoin (PTZ + PHT), 80 mg/kg topiramate (PTZ + TPR), 40 mg/kg lamotrigine (PTZ + LMT) and 50 mg/kg sodium valproate (PTZ + SV), respectively. All anticonvulsant drugs were injected 30 min prior to PTZ injection throughout 14 days. Following treatment period, behavioral, biochemical and immunohistochemical studies were performed.Results: PTZ + saline group revealed significantly decreased galanin, NPY, BDNF and NGF levels compared to control. PTZ + MDZ group had significantly increased galanin, BDNF and NGF levels compared to saline group. Also, PTZ + LEV group showed increased BDNF levels. PTZ + saline group revealed significantly lower neuron count and higher GFAP (+) cells in hippocampal CA1-CA3 regions. All anticonvulsants significantly reduced hippocampal astrogliosis whereas only midazolam, levetiracetam, sodium valproate and lamotrigine prevented neuronal loss.Conclusion: Our results suggested that anticonvulsant drugs may reduce the severity of seizures, and exert neuroprotective effects by altering the expression of neuropeptides and neurotrophins in the epileptogenic hippocampus.

Suppression of hTAS2R16 Signaling by Umami Substances.

Interaction between umami and bitter taste has long been observed in human sensory studies and in neural responses in animal models, however, the molecular mechanism for their action has not been delineated. Humans detect diverse bitter compounds using 25-30 members of the type 2 taste receptor (TAS2R) family of G protein-coupled receptor. In this study, we investigated the putative mechanism of antagonism by umami substances using HEK293T cells expressing hTAS2R16 and two known probenecid-insensitive mutant receptors, hTAS2R16 N96T and P44T. In wild type receptor, Glu-Glu, inosine monophosphate (IMP), and l-theanine behave as partial insurmountable antagonists, and monosodium glutamate (MSG) acts as a surmountable antagonist in comparison with probenecid as a full insurmountable antagonist. The synergism with IMP of umami substances still stands in the suppression of hTAS2R16 signaling. In mutagenesis analysis, we found that Glu-Glu, MSG, and l-theanine share at least one critical binding site on N96 and P44 with probenecid. These results provide the first evidence for a direct binding of umami substances to the hTAS2R16 through the probenecid binding pocket on the receptor, resulting in the suppression of bitterness.

Effects of inosine 5'-monophosphate supplementation in high fishmeal and high soybean diets on growth, immune-related gene expression in gibel carp (Carassius auratus gibelio var. CAS â ¢), and its challenge against Aeromonas hydrophila infection.

The present study was conducted to evaluate dietary inosine 5'-monophosphate (5'-IMP) on growth, immune genes expression and disease resistance against Aeromonas hydrophila in juvenile gibel carp (Carassius auratus gibelio var. CAS Ⅲ) (initial body weight: 7.48 g). Six diets were formulated containing exogenous 5'-IMP at three gradient levels (0, 0.1% and 0.2%) in the high dietary fishmeal group (15% fishmeal: D1, D2, D3) and in the high dietary soybean meal group (33% soybean meal: D4, D5, D6). Each diet was randomly allotted to triplicate tanks in a recirculating system. After the feeding trial, fish were exposed to Aeromonas hydrophila challenge. Hematological and immunological responses were analyzed before and after challenge. The results indicated that feeding rate in all 5'-IMP supplemented treatments (D2, D3, D5 and D6) and daily growth coefficient in D5 and D6 were reduced compared with those of respective control treatments (D1 and D4) without 5'-IMP addition (P < 0.05). The cumulative survival rates were numerically improved by dietary 5'-IMP supplementation (P > 0.05). Compared with the respective control treatment, in the high fishmeal group, plasma SOD and MPO were significantly elevated in D3 at the end of feeding trial (P < 0.05), plasma SOD and lysozyme were significantly increased in D3 after bacterial challenge (P < 0.05); in high soybean meal group, plasma lysozyme activity was significantly elevated in D5 post bacterial challenge (P < 0.05). Most of the expression of immune related genes (intelectin, major histocompatibility complex class II ß (MHC II ß), Complement 3 (C3), Complement component C7-1 (ccC7), lysozyme C, Interleukin 1ß (IL-1ß), Tumor necrosis factor α1 (TNF-α1), Transforming growth factor-beta (TGF-ß) and Interleukin 8 (IL-8)) in spleen, kidney and liver of the fish were significantly affected by supplementation of 5'-IMP at the end of feeding trial and post bacterial challenge. Additionally, adding 5'-IMP in high soybean meal diets exerted further effects of promoting immunity than counterparts in high fishmeal diets. Considering enhanced disease resistance, the immunopotentiation of 5'-IMP was manifested when the addition level was 0.1% in high soybean meal diets and 0.2% in high fishmeal diets.

Structure of Arabidopsis thaliana N6-methyl-AMP deaminase ADAL with bound GMP and IMP and implications for N6-methyl-AMP recognition and processing.

Arabidopsis thaliana aminohydrolase (AtADAL) has been shown to be involved in the metabolism of N6-methyl-AMP, a proposed intermediate during m6A-modified RNA metabolism, which can be subsequently incorporated into newly synthesized RNA by Pol II. It has been proposed that AtADAL will prevent N6-methyl-AMP reuse and catabolize it to inosine monophosphate (IMP). Here, we have solved the crystal structures of AtADAL in the apo form and in complex with GMP and IMP in the presence of Zn2+. We have identified the substrate-binding pocket of AtADAL and compared it with that for adenosine deaminase (ADA), adenine deaminase (ADE) and AMP deaminase (AMPD) from multiple species. The comparisons reveal that plant ADAL1 may have the potential ability to catalyze different alkyl-group substituted substrates.

Measuring deaminated nucleotide surveillance enzyme ITPA activity with an ATP-releasing nucleotide chimera.

Nucleotide quality surveillance enzymes play important roles in human health, by detecting damaged molecules in the nucleotide pool and deactivating them before they are incorporated into chromosomal DNA or adversely affect metabolism. In particular, deamination of adenine moiety in (deoxy)nucleoside triphosphates, resulting in formation of (d)ITP, can be deleterious, leading to DNA damage, mutagenesis and other harmful cellular effects. The 21.5 kDa human enzyme that mitigates this damage by conversion of (d)ITP to monophosphate, ITPA, has been proposed as a possible therapeutic and diagnostic target for multiple diseases. Measuring the activity of this enzyme is useful both in basic research and in clinical applications involving this pathway, but current methods are nonselective and are not applicable to measurement of the enzyme from cells or tissues. Here, we describe the design and synthesis of an ITPA-specific chimeric dinucleotide (DIAL) that replaces the pyrophosphate leaving group of the native substrate with adenosine triphosphate, enabling sensitive detection via luciferase luminescence signaling. The probe is shown to function sensitively and selectively to quantify enzyme activity in vitro, and can be used to measure the activity of ITPA in bacterial, yeast and human cell lysates.

Enterococcus faecium NCIMB 10415 supplementation improves the meat quality and antioxidant capacity of muscle of broilers.

The present study was conducted to investigate the effects of Enterococcus faecium (E. faecium) on the meat quality and antioxidant capacity of muscle in broilers. A total of 600 Arbor Acre broiler chickens (1-day-old, male) were randomly divided into five treatments with six replicates (20 chickens per replicate) for each treatment. The five treatments were the control treatment (CON, basal diet), antibiotic treatment (ANT, basal diet supplemented with 0.1% chlortetracycline) and E. faecium-supplemented treatments (LEF, MEF and HEF, basal diet supplemented with 50, 100 and 200 mg/kg of E. faecium respectively). The experiment lasted 42 days in two periods of 21 days. Results showed that there were no differences in breast meat quality among different treatments (p > 0.05). Compared with the CON and ANT treatments, the yellowness of thigh meat in E. faecium-supplemented treatments was significantly increased (p < 0.05); the shear force of thigh meat in the LEF and MEF treatments was lower than that of the CON treatment (p < 0.05). In addition, the concentration of the inosine monophosphate (IMP) in the breast and thigh meat of the MEF treatment was significantly higher than that of the other treatments (p < 0.05). At 21 days, the total antioxidant capability (T-AOC) level and glutathione peroxidase (GSH-Px) activity of breast meat and superoxide dismutase (SOD) in the thigh meat of the MEF treatment were greatly increased (p < 0.05). At 42 days of age, the catalase (CAT), GSH-Px and T-AOC activities in the breast meat of the MEF treatment were increased (p < 0.05) and the CAT activity of thigh meat in the LEF and MEF treatments was increased (p < 0.05). In conclusion, E. faecium supplementation increased the meat quality of the thigh muscle, increased the IMP content and the activities of CAT, SOD, T-AOC, and GSH-Px of muscle in broilers. Supplementation with 100 mg/kg E. faecium had the greatest effects.