RESUMEN
Long-term exposure to fine particulate matter (PM2.5) posed injury for gastrointestinal and respiratory systems, ascribing with the lung-gut axis. However, the cross-talk mechanisms remain unclear. Here, we attempted to establish the response networks of lung-gut axis in mice exposed to PM2.5 at environmental levels. Male Balb/c mice were exposed to PM2.5 (dose of 0.1, 0.5, and 1.0 mg/kg) collected from Chengdu, China for 10 weeks, through intratracheally instillation, and examined the effect of PM2.5 on lung functions of mice. The changes of lung and gut microbiota and metabolic profiles of mice in different groups were determined. Furthermore, the results of multi-omics were conjointly analyzed to elucidate the primary microbes and the associated metabolites in lung and gut responsible for PM2.5 exposure. Accordingly, the cross-talk network and key pathways between lung-gut axis were established. The results indicated that exposed to PM2.5 0.1 mg/kg induced obvious inflammations in mice lung, while emphysema was observed at 1.0 mg/kg. The levels of metabolites guanosine, hypoxanthine, and hepoxilin B3 increased in the lung might contribute to lung inflammations in exposure groups. For microbiotas in lung, PM2.5 exposure significantly declined the proportions of Halomonas and Lactobacillus. Meanwhile, the metabolites in gut including L-tryptophan, serotonin, and spermidine were up-regulated in exposure groups, which were linked to the decreasing of Oscillospira and Helicobacter in gut. Via lung-gut axis, the activations of pathways including Tryptophan metabolism, ABC transporters, Serotonergic synapse, and Linoleic acid metabolism contributed to the cross-talk between lung and gut tissues of mice mediated by PM2.5. In summary, the microbes including Lactobacillus, Oscillospira, and Parabacteroides, and metabolites including hepoxilin B3, guanosine, hypoxanthine, L-tryptophan, and spermidine were the main drivers. In this lung-gut axis study, we elucidated some pro- and pre-biotics in lung and gut microenvironments contributed to the adverse effects on lung functions induced by PM2.5 exposure.
Asunto(s)
Contaminantes Atmosféricos , Lesión Pulmonar , Masculino , Ratones , Animales , Lesión Pulmonar/inducido químicamente , Contaminantes Atmosféricos/toxicidad , Contaminantes Atmosféricos/metabolismo , Triptófano , Multiómica , Espermidina/metabolismo , Espermidina/farmacología , Pulmón , Material Particulado/toxicidad , Material Particulado/metabolismo , Guanosina/metabolismo , Guanosina/farmacología , Hipoxantinas/metabolismo , Hipoxantinas/farmacologíaRESUMEN
OBJECTIVES: To investigate the effect of the L-arginine metabolism on arthritis and inflammation-mediated bone loss. METHODS: L-arginine was applied to three arthritis models (collagen-induced arthritis, serum-induced arthritis and human TNF transgenic mice). Inflammation was assessed clinically and histologically, while bone changes were quantified by µCT and histomorphometry. In vitro, effects of L-arginine on osteoclast differentiation were analysed by RNA-seq and mass spectrometry (MS). Seahorse, Single Cell ENergetIc metabolism by profilIng Translation inHibition and transmission electron microscopy were used for detecting metabolic changes in osteoclasts. Moreover, arginine-associated metabolites were measured in the serum of rheumatoid arthritis (RA) and pre-RA patients. RESULTS: L-arginine inhibited arthritis and bone loss in all three models and directly blocked TNFα-induced murine and human osteoclastogenesis. RNA-seq and MS analyses indicated that L-arginine switched glycolysis to oxidative phosphorylation in inflammatory osteoclasts leading to increased ATP production, purine metabolism and elevated inosine and hypoxanthine levels. Adenosine deaminase inhibitors blocking inosine and hypoxanthine production abolished the inhibition of L-arginine on osteoclastogenesis in vitro and in vivo. Altered arginine levels were also found in RA and pre-RA patients. CONCLUSION: Our study demonstrated that L-arginine ameliorates arthritis and bone erosion through metabolic reprogramming and perturbation of purine metabolism in osteoclasts.
Asunto(s)
Artritis Experimental , Artritis Reumatoide , Resorción Ósea , Humanos , Ratones , Animales , Osteoclastos , Artritis Reumatoide/patología , Artritis Experimental/patología , Inflamación/metabolismo , Ratones Transgénicos , Arginina/farmacología , Inosina/metabolismo , Inosina/farmacología , Hipoxantinas/metabolismo , Hipoxantinas/farmacología , Purinas/farmacologíaRESUMEN
The present study investigated the role of heat shock protein 90 (HSP90) in the proliferation and survival of Babesia gibsoni in vitro. To detect the effect on the entry of B. gibsoni into host erythrocytes, the parasite was incubated with an antibody against B. gibsoni HSP90 (BgHSP90) for 24 h. The results of this experiment demonstrated that both the incorporation of [3H]hypoxanthine into the nucleic acids of B. gibsoni and the number of parasites were not altered, indicating that an anti-BgHSP90 antibody did not directly inhibit the entry of the parasite into erythrocytes. Moreover, two HSP90 inhibitors, geldanamycin (GA) and tanespimycin (17-AAG), were used to evaluate the function of BgHSP90. GA and 17-AAG decreased both the incorporation of [3H]hypoxanthine and the number of infected erythrocytes, suggesting that BgHSP90 plays important roles in DNA synthesis and the proliferation of B. gibsoni. The effect of 17-AAG on the parasites was weaker than that of GA. Additionally, the effect of GA on the survival and superoxide generation of canine neutrophils was assessed. The survival of canine neutrophils was not affected. The superoxide generation was strongly suppressed by GA. This result indicated that GA inhibited the function of canine neutrophils. Additional studies are necessary to elucidate the role of BgHSP90 in the proliferation of the parasite.
Asunto(s)
Babesia , Babesiosis , Enfermedades de los Perros , Animales , Perros , Superóxidos/metabolismo , Proteínas HSP90 de Choque Térmico/metabolismo , Hipoxantinas/metabolismo , Hipoxantinas/farmacología , Proliferación Celular , Enfermedades de los Perros/parasitología , Babesiosis/parasitologíaRESUMEN
BACKGROUND: Lowered nicotinamide adenine dinucleotide (NAD+) levels in tumor cells drive tumor hyperprogression during immunotherapy, and its restoration activates immune cells. However, the effect of lenvatinib, a first-line treatment for unresectable hepatocellular carcinoma (HCC), on NAD+ metabolism in HCC cells, and the metabolite crosstalk between HCC and immune cells after targeting NAD+ metabolism of HCC cells remain unelucidated. METHODS: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) and ultra-high-performance liquid chromatography multiple reaction monitoring-mass spectrometry (UHPLC-MRM-MS) were used to detect and validate differential metabolites. RNA sequencing was used to explore mRNA expression in macrophages and HCC cells. HCC mouse models were used to validate the effects of lenvatinib on immune cells and NAD+ metabolism. The macrophage properties were elucidated using cell proliferation, apoptosis, and co-culture assays. In silico structural analysis and interaction assays were used to determine whether lenvatinib targets tet methylcytosine dioxygenase 2 (TET2). Flow cytometry was performed to assess changes in immune cells. RESULTS: Lenvatinib targeted TET2 to synthesize and increase NAD+ levels, thereby inhibiting decomposition in HCC cells. NAD+ salvage increased lenvatinib-induced apoptosis of HCC cells. Lenvatinib also induced CD8+ T cells and M1 macrophages infiltration in vivo. And lenvatinib suppressed niacinamide, 5-Hydroxy-L-tryptophan and quinoline secretion of HCC cells, and increased hypoxanthine secretion, which contributed to proliferation, migration and polarization function of macrophages. Consequently, lenvatinib targeted NAD+ metabolism and elevated HCC-derived hypoxanthine to enhance the macrophages polarization from M2 to M1. Glycosaminoglycan binding disorder and positive regulation of cytosolic calcium ion concentration were characteristic features of the reverse polarization. CONCLUSIONS: Targeting HCC cells NAD+ metabolism by lenvatinib-TET2 pathway drives metabolite crosstalk, leading to M2 macrophages reverse polarization, thereby suppressing HCC progression. Collectively, these novel insights highlight the role of lenvatinib or its combination therapies as promising therapeutic alternatives for HCC patients with low NAD+ levels or high TET2 levels.
Asunto(s)
Carcinoma Hepatocelular , Neoplasias Hepáticas , Quinolinas , Animales , Ratones , Humanos , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , NAD/metabolismo , NAD/farmacología , NAD/uso terapéutico , Linfocitos T CD8-positivos , Cromatografía Liquida , Línea Celular Tumoral , Espectrometría de Masas en Tándem , Macrófagos/metabolismo , Quinolinas/farmacología , Quinolinas/uso terapéutico , Hipoxantinas/metabolismo , Hipoxantinas/farmacología , Hipoxantinas/uso terapéuticoRESUMEN
2-Azahypoxanthine (AHX) and 2-aza-8-oxohypoxanthine (AOH), discovered as causal substances of fairy rings are known to be endogenous in the fairy ring-forming Lepista sordida. In this study, we showed that xanthine dioxygenase, an a-ketoglutarate-dependent dioxygenase, might catalyze the conversion of AHX to AOH in the fungus. Furthermore, this enzyme is the first reported molybdopterin-independent protein of hypoxanthine metabolism.
Asunto(s)
Agaricales , Dioxigenasas , Vías Biosintéticas , Xantina/metabolismo , Dioxigenasas/metabolismo , Agaricales/metabolismo , Hipoxantinas/metabolismoRESUMEN
2-Azahypoxanthine was isolated from the fairy ring-forming fungus Lepista sordida as a fairy ring-inducing compound. 2-Azahypoxanthine has an unprecedented 1,2,3-triazine moiety, and its biosynthetic pathway is unknown. The biosynthetic genes for 2-azahypoxanthine formation in L. sordida were predicted by a differential gene expression analysis using MiSeq. The results revealed that several genes in the purine and histidine metabolic pathways and the arginine biosynthetic pathway are involved in the biosynthesis of 2-azahypoxanthine. Furthermore, nitric oxide (NO) was produced by recombinant NO synthase 5 (rNOS5), suggesting that NOS5 can be the enzyme involved in the formation of 1,2,3-triazine. The gene encoding hypoxanthine-guanine phosphoribosyltransferase (HGPRT), one of the major phosphoribosyltransferases of purine metabolism, increased when 2-azahypoxanthine content was the highest. Therefore, we hypothesized that HGPRT might catalyze a reversible reaction between 2-azahypoxanthine and 2-azahypoxanthine-ribonucleotide. We proved the endogenous existence of 2-azahypoxanthine-ribonucleotide in L. sordida mycelia by LC-MS/MS for the first time. Furthermore, it was shown that recombinant HGPRT catalyzed reversible interconversion between 2-azahypoxanthine and 2-azahypoxanthine-ribonucleotide. These findings demonstrate that HGPRT can be involved in the biosynthesis of 2-azahypoxanthine via 2-azahypoxanthine-ribonucleotide generated by NOS5.
Asunto(s)
Agaricales , Hipoxantina Fosforribosiltransferasa , Hipoxantina Fosforribosiltransferasa/genética , Hipoxantina Fosforribosiltransferasa/metabolismo , Cromatografía Liquida , Transcriptoma , Espectrometría de Masas en Tándem , Agaricales/metabolismo , Hipoxantinas/metabolismo , Ribonucleótidos/metabolismoRESUMEN
Hypoxanthine is a purine metabolite which increases during hypoxia and therefore is an indicator of this condition. Further, when hypoxanthine is oxidized to uric acid in the presence of xanthine oxidase, oxygen radicals are generated. This was the theoretical basis for suggesting and studying, beginning in the 1990s, resuscitation of newborn infants with air instead of the traditional 100% O2. These studies demonstrated a 30% reduction in mortality when resuscitation of term and near term infants was carried out with air compared to pure oxygen. The mechanism for this is not fully understood, however the hypoxanthine -xanthine oxidase system increases oxidative stress and plays a role in regulation of the perinatal circulation. Further, hyperoxic resuscitation inhibits mitochondrial function, and one reason may be that genes involved in ATP production are down-regulated. Thus, the study of one single molecule, hypoxanthine, has contributed to the global prevention of an estimated 2-500,000 annual infant deaths.
Asunto(s)
Hipoxantina , Hipoxia , Oxígeno , Femenino , Humanos , Lactante , Recién Nacido , Embarazo , Hipoxantina/metabolismo , Hipoxantinas/metabolismo , Hipoxia/metabolismo , Oxígeno/metabolismo , Ácido Úrico/metabolismo , Xantina Oxidasa/metabolismoRESUMEN
SCOPE: This study aims to investigate the anti-hyperuricemic and nephroprotective effects and the potential mechanisms of the separated gastrointestinal hydrolysates of α-lactalbumin on hyperuricemic mice. METHODS AND RESULTS: The gastrointestinal hydrolysate of α-lactalbumin, the hydrolysate fraction with molecular weight (MW) < 3 kDa (LH-3k), and the fragments with smallest MW among LH-3K harvested through dextran gel chromatography (F5) are used. Hyperuricemia mice are induced via daily oral gavage of potassium oxonate and hypoxanthine. F5 displays the highest in vitro xanthine oxidase (XO) inhibition among all the fractions separated from LH-3k. Oral administration of F5 significantly reduces the levels of serum uric acid (UA), creatinine, and urea nitrogen. F5 treatment could ameliorate kidney injury through alleviating oxidative stress and inflammation. F5 alleviates hyperuricemia in mice by inhibiting hepatic XO activity and regulating the expression of renal urate transporters. Gut microbiota analysis illustrates that F5 administration increases the abundance of some SCFAs producers, and inhibits the growth of hyperuricemia and inflammation associated genera. LH-3k exhibits similar effects but does not show significance as those of the F5 fraction. CONCLUSION: The anti-hyperuricemia and nephroprotective functions of F5 are mediated by inhibiting hepatic XO activity, ameliorating oxidative stress and inflammation, regulating renal urate transporters, and modulating the gut microbiota in hyperuricemic mice.
Asunto(s)
Microbioma Gastrointestinal , Hiperuricemia , Ratones , Animales , Ácido Úrico , Lactalbúmina/metabolismo , Hiperuricemia/tratamiento farmacológico , Riñón/metabolismo , Ácido Oxónico/metabolismo , Ácido Oxónico/farmacología , Factores de Transcripción/metabolismo , Inflamación/metabolismo , Hipoxantinas/metabolismo , Hipoxantinas/farmacologíaRESUMEN
OBJECTIVE: To investigate the protective effect and mechanism of polydatin (PD) against gouty nephropathy (GN) in mice. METHODS: Twenty-four mice were randomly divided into three groups: the control group (no treatment), the GN group (300 mg/kg hypoxanthine + 150 mg/kg potassium oxonate), and the GN + PD group (300 mg/kg hypoxanthine + 150 mg/kg potassium oxonate + 50 mg/kg PD). Histological changes in the kidneys and the levels of uric acid (UA), blood urea nitrogen (BUN), and serum creatinine (SCr) in the sera were measured. In addition, the expression of gasdermin D (GSDMD) protein in renal tubular epithelial cells, and the expression of NOD-like receptor protein 3 (NLRP3), GSDMD, and caspase-1 proteins in the kidney tissues were determined by immunohistochemistry, immunofluorescence, and Western blot. RESULTS: In vitro, PD inhibited the expression of NLRP3, caspase-1, and GSDMD and protected the renal tubular epithelial cells from pyroptosis. In vivo, PD treatment significantly ameliorated the pathological changes in kidney tissue, and reversed the decrease of serum UA and BUN in GN model mice. The expression of NLRP3, GSDMD, and caspase-1 proteins was also decreased in the PD-treated GN mice. CONCLUSION: The results suggest that PD has a protective effect on mice with GN, which may be related to the downregulation of NLRP3, GSDMD, and caspase-1 proteins and the inhibition of renal tubular epithelial cells pyroptosis.
Asunto(s)
Gota , Proteína con Dominio Pirina 3 de la Familia NLR , Animales , Ratones , Caspasas , Células Epiteliales/metabolismo , Gota/metabolismo , Hipoxantinas/metabolismo , Hipoxantinas/farmacología , Riñón/patología , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Piroptosis/fisiologíaRESUMEN
In both animals and higher plants, xanthine dehydrogenase is a highly conserved housekeeping enzyme in purine degradation where it oxidizes hypoxanthine to xanthine and xanthine to uric acid. Previous reports demonstrated that xanthine dehydrogenase played a vital role in N metabolism and stress response. Is xanthine dehydrogenase involved in regulating leaf senescence? A recessive early senescence mutant with excess sugar accumulation, ossac3, was isolated previously by screening the EMS-induced mutant library. Here, we show that xanthine dehydrogenase not only plays a role in N metabolism but also involved in regulating carbon metabolism in rice. Based on map-based cloning, OsSAC3 was identified, which encodes the xanthine dehydrogenase. OsSAC3 was constitutively expressed in all examined tissues and the OsSAC3 protein located in the cytoplasm. Transcriptional analysis revealed purine metabolism, chlorophyll metabolism, photosynthesis, sugar metabolism and redox balance were affected in the ossac3 mutant. Moreover, carbohydrate distribution was changed, leading to the accumulation of sucrose and starch in the leaves containing ossac3 on account of decreased expression of OsSWEET3a, OsSWEET6a and OsSWEET14 and oxidized inactivation of starch degradation enzymes in ossac3. These results indicated that OsSAC3 played a vital role in leaf senescence by regulating carbon metabolism in rice.
Asunto(s)
Oryza , Carbohidratos , Carbono/metabolismo , Clorofila/metabolismo , Regulación de la Expresión Génica de las Plantas , Hipoxantinas/metabolismo , Mutación , Oryza/fisiología , Fenotipo , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Almidón/metabolismo , Sacarosa/metabolismo , Azúcares/metabolismo , Ácido Úrico/metabolismo , Xantina Deshidrogenasa/genética , Xantina Deshidrogenasa/metabolismoRESUMEN
Accumulated clinical and biomedical evidence indicates that the gut microbiota and their metabolites affect brain function and behavior in various central nervous system disorders. This study was performed to investigate the changes in brain metabolites and composition of the fecal microbial community following injection of amyloid ß (Aß) and donepezil treatment of Aß-injected mice using metataxonomics and metabolomics. Aß treatment caused cognitive dysfunction, while donepezil resulted in the successful recovery of memory impairment. The Aß + donepezil group showed a significantly higher relative abundance of Verrucomicrobia than the Aß group. The relative abundance of 12 taxa, including Blautia and Akkermansia, differed significantly between the groups. The Aß + donepezil group had higher levels of oxalate, glycerol, xylose, and palmitoleate in feces and oxalate, pyroglutamic acid, hypoxanthine, and inosine in brain tissues than the Aß group. The levels of pyroglutamic acid, glutamic acid, and phenylalanine showed similar changes in vivo and in vitro using HT-22 cells. The major metabolic pathways in the brain tissues and gut microbiota affected by Aß or donepezil treatment of Aß-injected mice were related to amino acid pathways and sugar metabolism, respectively. These findings suggest that alterations in the gut microbiota might influence the induction and amelioration of Aß-induced cognitive dysfunction via the gut-brain axis. This study could provide basic data on the effects of Aß and donepezil on gut microbiota and metabolites in an Aß-induced cognitive impairment mouse model.
Asunto(s)
Enfermedad de Alzheimer , Disfunción Cognitiva , Microbioma Gastrointestinal , Enfermedad de Alzheimer/inducido químicamente , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/metabolismo , Animales , Encéfalo/metabolismo , Disfunción Cognitiva/inducido químicamente , Disfunción Cognitiva/tratamiento farmacológico , Disfunción Cognitiva/metabolismo , Modelos Animales de Enfermedad , Donepezilo/farmacología , Donepezilo/uso terapéutico , Ácido Glutámico/metabolismo , Glicerol/metabolismo , Hipoxantinas/metabolismo , Hipoxantinas/farmacología , Hipoxantinas/uso terapéutico , Inosina/metabolismo , Ratones , Oxalatos/metabolismo , Fenilalanina/metabolismo , Ácido Pirrolidona Carboxílico/metabolismo , Xilosa/metabolismoRESUMEN
This study aims to investigate the anti-hyperuricemia effect and mechanism of anserine in hyperuricemic rats. Hyperuricemic rats were induced with a combination of 750 mg per kg bw d potassium oxazinate (PO) and 200 mg per kg bw d hypoxanthine for a week, and the rats were separately orally administered anserine (20, 40, 80 mg kg-1) and allopurinol (10 mg kg-1) for three weeks. The results show that the content of serum uric acid (SUA) decreased by approximately 40% and 60% after the intervention of anserine and allopurinol, respectively. The activity of superoxide dismutase (SOD) was increased and the levels of malondialdehyde (MDA), alkaline phosphatase (ALP) and alanine aminotransferase (ALT) were significantly decreased in the anserine groups. After the administration of anserine, the contents of blood urea nitrogen (BUN) and creatinine (Cr) were reduced in the kidney, and the levels of the proinflammatory cytokines IL-1ß, IL-6ß, TNF-α and TGF-ß and inflammatory cell infiltration were reduced in both the liver and kidney. Moreover, the gene expressions of xanthine oxidase (XOD), renal urate transporter 1 (URAT1) and glucose transporter type 9 (GLUT9) were downregulated by anserine administration, and the gene expressions of ATP-binding cassette transporter G2 (ABCG2), organic anion transporter 1 (OAT1) and organic anion transporter 3 (OAT3) were upregulated at the same time. These findings suggest that hepatorenal injury was repaired by anserine, which further regulated the expression of hepatic XOD and renal URAT1, GLUT9, ABCG2, OAT1 and OAT3 to relieve hyperuricemia in rats.
Asunto(s)
Hiperuricemia , Transportadores de Anión Orgánico , Transportadoras de Casetes de Unión a ATP/metabolismo , Alanina Transaminasa/metabolismo , Fosfatasa Alcalina/metabolismo , Alopurinol/metabolismo , Alopurinol/farmacología , Animales , Anserina/metabolismo , Anserina/farmacología , Creatinina , Proteínas Facilitadoras del Transporte de la Glucosa/metabolismo , Hiperuricemia/metabolismo , Hipoxantinas/metabolismo , Hipoxantinas/farmacología , Riñón , Hígado/metabolismo , Malondialdehído/metabolismo , Transportadores de Anión Orgánico/genética , Transportadores de Anión Orgánico/metabolismo , Potasio/metabolismo , Potasio/farmacología , Ratas , Superóxido Dismutasa/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Ácido Úrico , Xantina Oxidasa/metabolismoRESUMEN
Among the "fairy chemicals" involved in forming the natural phenomenon of "fairy rings," we focused on 2-aza-8-oxohypoxanthine (AOH) as a candidate functional cosmetic ingredient. In previous studies, AOH was confirmed to be safe for use on human skin, and no adverse reactions were observed in any of the safety studies. In this study, we report the results of a clinical trial using a lotion containing AOH. Our analysis using the L* value for indices of skin lightness indicated that the AOH application significantly increased the L* value after 8 weeks. Since a previous DNA microarray study using normal human epidermal cells showed that AOH suppressed the expression of a group of genes that induce inflammatory cytokines (prostaglandin E synthase, prostaglandin-endoperoxide synthase 2 [cyclooxygenase-2], and interleukin-18), our results suggest that the AOH-induced suppression of inflammatory factors results in skin lightening.
Asunto(s)
Cosméticos , Ciclooxigenasa 2/genética , Humanos , Hipoxantinas/metabolismo , Piel/metabolismoRESUMEN
Helicobacter pylori (Hp) is a human pathogen that lives in the gastric mucosa of approximately 50% of the world's population causing gastritis, peptic ulcers, and gastric cancer. An increase in resistance to current drugs has sparked the search for new Hp drug targets and therapeutics. One target is the disruption of nucleic acid production, which can be achieved by impeding the synthesis of 6-oxopurine nucleoside monophosphates, the precursors of DNA and RNA. These metabolites are synthesized by Hp xanthine-guanine-hypoxanthine phosphoribosyltransferase (XGHPRT). Here, nucleoside phosphonates have been evaluated, which inhibit the activity of this enzyme with Ki values as low as 200 nM. The prodrugs of these compounds arrest the growth of Hp at a concentration of 50 µM in cell-based assays. The kinetic properties of HpXGHPRT have been determined together with its X-ray crystal structure in the absence and presence of 9-[(N-3-phosphonopropyl)-aminomethyl-9-deazahypoxanthine, providing a basis for new antibiotic development.
Asunto(s)
Antibacterianos/química , Proteínas Bacterianas/metabolismo , Pentosiltransferasa/metabolismo , Secuencia de Aminoácidos , Antibacterianos/metabolismo , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Proteínas Bacterianas/química , Sitios de Unión , Cristalografía por Rayos X , Enfermedades Gastrointestinales/tratamiento farmacológico , Enfermedades Gastrointestinales/microbiología , Enfermedades Gastrointestinales/patología , Infecciones por Helicobacter/tratamiento farmacológico , Infecciones por Helicobacter/patología , Helicobacter pylori/efectos de los fármacos , Helicobacter pylori/enzimología , Humanos , Hipoxantina Fosforribosiltransferasa/química , Hipoxantina Fosforribosiltransferasa/metabolismo , Hipoxantinas/química , Hipoxantinas/metabolismo , Hipoxantinas/farmacología , Hipoxantinas/uso terapéutico , Cinética , Simulación de Dinámica Molecular , Organofosfonatos/química , Organofosfonatos/metabolismo , Organofosfonatos/farmacología , Organofosfonatos/uso terapéutico , Pentosiltransferasa/química , Profármacos/química , Profármacos/metabolismo , Profármacos/farmacología , Profármacos/uso terapéutico , Alineación de Secuencia , Relación Estructura-ActividadRESUMEN
Fairy rings resulting from a fungus-plant interaction appear worldwide. 2-Azahypoxanthine (AHX) and imidazole-4-carboxamide (ICA) were first isolated from the culture broth of one of the fairy ring-forming fungi, Lepista sordida. Afterward, a common metabolite of AHX in plants, 2-aza-8-oxohypoxanthine (AOH), was found in AHX-treated rice. The biosynthetic pathway of the three compounds that are named as fairy chemicals (FCs) in plants has been partially elucidated; however, that in mushrooms remains unknown. In this study, it was revealed that the carbon skeletons of AHX and ICA were constructed from Gly in L. sordida mycelia and the fungus metabolized 5-aminoimidazole-4-carboxamide (AICA) to both of the compounds. These results indicated that FCs were biosynthesized by a diversion of the purine metabolic pathway in L. sordida mycelia, similar to that in plants. Furthermore, we showed that recombinant adenine phosphoribosyltransferase (APRT) catalyzed reversible interconversion not only between 5-aminoimidazole-4-carboxamide-1-ß-d-ribofuranosyl 5'-monophosphate (AICAR) and AICA but also between ICA-ribotide (ICAR) and ICA. Furthermore, the presence of ICAR in L. sordida mycelia was proven for the first time by LC-MS/MS detection, and this study provided the first report that there was a novel metabolic pathway of ICA in which its ribotide was an intermediate in the fungus.
Asunto(s)
Agaricales/metabolismo , Aminoimidazol Carboxamida/análogos & derivados , Hipoxantinas/metabolismo , Aminoimidazol Carboxamida/química , Aminoimidazol Carboxamida/aislamiento & purificación , Aminoimidazol Carboxamida/metabolismo , Espectroscopía de Resonancia Magnética con Carbono-13 , Cromatografía Liquida , Cuerpos Fructíferos de los Hongos/química , Hipoxantinas/química , Hipoxantinas/aislamiento & purificación , Espectrometría de Masas en TándemRESUMEN
Circular patterns called "fairy rings" in fields are a natural phenomenon that arises through the interaction between basidiomycete fungi and plants. Acceleration or inhibition of plant vegetative growth and the formation of mushroom fruiting bodies are both commonly observed when fairy rings form. The gene of an enzyme involved in the biosynthesis of these regulators was recently isolated in the fairy ring-forming fungus, Lepista sordida. To identify other genes involved in L. sordida fairy ring formation, we used previously generated sequence data to produce a more complete draft genome sequence for this species. Finally, we predicted the metabolic pathways of the plant growth regulators and 29 candidate enzyme-coding genes involved in fairy-ring formation based on gene annotations. Comparisons of protein coding genes among basidiomycete fungi revealed two nitric oxide synthase gene candidates that were uniquely encoded in genomes of fairy ring-forming fungi. These results provide a basis for the discovery of genes involved in fairy ring formation and for understanding the mechanisms involved in the interaction between fungi and plants. We also constructed a new web database F-RINGS ( http://bioinf.mind.meiji.ac.jp/f-rings/ ) to provide the comprehensive genomic information for L. sordida.
Asunto(s)
Agaricales/genética , Cuerpos Fructíferos de los Hongos/genética , Genoma Fúngico , Plantas/microbiología , Agaricales/enzimología , Bases de Datos Genéticas , Enzimas/química , Enzimas/genética , Cuerpos Fructíferos de los Hongos/metabolismo , Hipoxantinas/metabolismo , Redes y Vías Metabólicas/genética , Óxido Nítrico Sintasa/química , Óxido Nítrico Sintasa/genética , Reguladores del Crecimiento de las Plantas/metabolismo , Plantas/genética , Plantas/metabolismo , Análisis de Secuencia de ADNRESUMEN
An Escherichia coli strain was constructed for the efficient import of nicotinamide adenine dinucleotide (NAD) analogues into cells by limiting extracellular degradation while expressing an efficient NAD importer. In vivo functions of three NAD analogues were characterized. Nicotinamide hypoxanthine dinucleotide was identified as an inhibitor of NAD synthesis. Nicotinamide cytosine dinucleotide had excellent biocompatibility and was used for characterizing a growth-dependent degradation of in vivo nicotinamide cofactors.
Asunto(s)
Escherichia coli/metabolismo , NAD/análogos & derivados , NAD/metabolismo , Niacinamida/química , Coenzimas/metabolismo , Citosina/análogos & derivados , Citosina/metabolismo , Citosina/farmacología , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Hipoxantinas/metabolismo , Hipoxantinas/farmacología , Estructura Molecular , Mutación , NAD/farmacología , Hidrolasas Diéster Fosfóricas/genética , Hidrolasas Diéster Fosfóricas/metabolismo , Pirofosfatasas/genética , Pirofosfatasas/metabolismoRESUMEN
"Fairy rings" resulting from fungus-stimulated plant growth occur all over the world. In 2010, 2-azahypoxanthine (AHX) from a fungus Lepista sordida was identified as the "fairy" that stimulates plant growth. Furthermore, 2-aza-8-oxohypoxanthine (AOH) was isolated as a common metabolite of AHX in plants, and the endogenous existence of AHX and AOH in plants was proved. The structure of AHX allowed us to hypothesize that AHX was derived from 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR). Thus, we performed a feeding experiment that supplied AICAR to L. sordida. Consumption of AICAR and accumulation of AHX were observed after feeding. The mycelia extract had enzymatic activity of adenine/5-aminoimidazole-4-carboxamide phosphoribosyltransferase (APRT). APRT gene of L. sordida revealed its structural characteristics in homology modeling and showed transcriptional enhancement after feeding. These results support that AHX was synthesized from AICAR and AHX biosynthesis was transcriptionally controlled by AICAR, indicating the presence of novel purine metabolic pathway in L. sordida.
Asunto(s)
Adenina Fosforribosiltransferasa/genética , Agaricales/enzimología , Hipoxantinas/metabolismo , Adenina Fosforribosiltransferasa/química , Adenina Fosforribosiltransferasa/metabolismo , Agaricales/genética , Agaricales/metabolismo , Aminoimidazol Carboxamida/análogos & derivados , Aminoimidazol Carboxamida/química , Aminoimidazol Carboxamida/metabolismo , Vías Biosintéticas , Clonación Molecular , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Hipoxantinas/química , Conformación Proteica , Ribonucleótidos/química , Ribonucleótidos/metabolismoRESUMEN
Fairy rings are zones of stimulated grass growth owing to the interaction between a fungus and a plant. We previously reported the discovery of two novel plant-growth regulating compounds related to forming fairy rings, 2-azahypoxanthine (AHX) and 2-aza-8-oxohypoxanthine (AOH). In this study, a bacterial strain CH-1 was isolated from an airborne-contaminated nutrient medium containing AHX. The strain converted AHX to AOH and identified as Burkholderia contaminans based on the gene sequence of its 16S rDNA. The quantitative production of AOH by resting cells of the strain was achieved. Among seven Burkholderia species, two bacteria and two yeasts tested, B. contaminans CH-1 showed the highest rate of conversion of AHX to AOH. By batch system, up to 10.6 mmol AHX was converted to AOH using the resting cells. The yield of this process reached at 91%.
Asunto(s)
Burkholderia/citología , Burkholderia/metabolismo , Hipoxantinas/metabolismo , Microbiología del Aire , Biotransformación , Burkholderia/aislamiento & purificaciónRESUMEN
Rings or arcs of fungus-stimulated plant growth occur worldwide; these are commonly referred to as "fairy rings". In 2010, we discovered 2-azahypoxanthine (AHX), a compound responsible for the fairy-ring phenomenon caused by fungus; AHX stimulated the growth of all the plants tested. Herein, we reveal the isolation and structure determination of a common metabolite of AHX in plants, 2-aza-8-oxohypoxanthine (AOH). AHX is chemically synthesized from 5-aminoimidazole-4-carboxamide (AICA), and AHX can be converted into AOH by xanthine oxidase. AICA is one of the members of the purine metabolic pathway in animals, plants, and microorganisms. However, further metabolism of AICA remains elusive. Based on these results and facts, we hypothesized that plants themselves produce AHX and AOH through a pathway similar to the chemical synthesis. Herein, we demonstrate the existence of endogenous AHX and AOH and a novel purine pathway to produce them in plants.