Background During the weaning phase,piglets are exposed to significant physiological and environmental stressors,which disrupt the balance of their intestinal microbiota and often lead to severe diarrhea.Previous stud...Background During the weaning phase,piglets are exposed to significant physiological and environmental stressors,which disrupt the balance of their intestinal microbiota and often lead to severe diarrhea.Previous studies have demonstrated that alfalfa fiber,derived from the stems and leaves of alfalfa,can effectively alleviate diarrhea in piglets.Additionally,multiple studies have highlighted the potential of fecal microbiota transplantation(FMT)in mitigating diarrhea in various models of intestinal diseases in young animals.However,the specific mechanisms by which FMT from targeted sources alleviates diarrhea in weaned piglets remain to be fully elucidated.Results In this study,FMT from donor piglets fed an alfalfa fiber-supplemented diet effectively alleviated diarrhea,improved intestinal morphology,and enhanced gut barrier function in weaned piglets.FMT further promoted the colonization of beneficial bacterial genera(including UCG-005,unclassified Lachnospiraceae,Lachnospiraceae AC2044 group,UCG-002,Candidatus Saccharimonas,and Lachnospiraceae ND3007 group)while inhibiting the detrimental genus Tyzzerella,consequently enhancing the production of short-chain fatty acids(SCFAs).Additionally,FMT upregulated riboflavin metabolism,leading to elevated flavin adenine dinucleotide(FAD)levels and increased glutathione reductase activity,thereby collectively attenuating lipopolysaccharide(LPS)-induced oxidative stress and contributing to intestinal health.Conclusions We found that FMT modulates the structure of the gut microbiota,enhances microbial diversity and composition,increases the production of SCFAs,and upregulates riboflavin metabolism to elevate FAD levels.These changes collectively enhance immune and antioxidant capacities,thereby alleviating diarrhea.展开更多
Microbes can cause or accelerate metal corrosion,leading to huge losses in corrosion damages each year.Geobacter sulfurreducens is a representative electroactive bacterium in many soils,sediments,and wastew-ater syste...Microbes can cause or accelerate metal corrosion,leading to huge losses in corrosion damages each year.Geobacter sulfurreducens is a representative electroactive bacterium in many soils,sediments,and wastew-ater systems.It has been confirmed to directly extract electrons from elemental metals.However,little is known about the effect of electron shuttles in G.sulfurreducens corrosion on stainless steel.In this study,we report that exogenous flavins promote iron-to-microbe electron transfer,accelerating micro-bial corrosion.G.sulfurreducens caused 1.3 times deeper pits and increased electron uptake(with 2 times increase of i_(corr))from stainless steel when riboflavin was added to the culture medium.OmcS-deficient mutant data suggest that G.sulfurreducens utilizes riboflavin as a bound-cofactor in outer membrane c-type cytochromes.The finding that,in the presence of microbes,riboflavin can substantially accelerate corrosion highlights the role of flavin redox cycling for enhanced iron-to-microbe electron transfer by G.sulfurreducens and provides new insights in microbial corrosion.展开更多
The aim of this study was to examine whether flavin-containing monooxygenase (FMO) protein was expressed in cultured rat brain microvascular endothelial cells (BMECs), which constitute the blood-brain barrier (BBB), a...The aim of this study was to examine whether flavin-containing monooxygenase (FMO) protein was expressed in cultured rat brain microvascular endothelial cells (BMECs), which constitute the blood-brain barrier (BBB), and whether N-oxide from the tertiary amine, d-chlorpheniramine, was formed by FMO in rat BMECs. BMECs were isolated and cultured from the brains of three-week-old male Wistar rats. The expression of FMO1, FMO2 and FMO5 proteins was confirmed in rat BMECs by western blotting analysis using polyclonal anti-FMO antibodies, but FMO3 and FMO4 proteins were not found in the rat BBB. Moreover, N-oxide of d-chlorpheniramine was formed in rat BMECs. The intrinsic clearance value for N-oxidation at pH 8.4 was higher than that at pH 7.4. Inhibition of N-oxide formation by methimazole was found to be the best model of competitive inhibition yielding an apparent Ki value of 0.53 μmol/L, suggesting that N-oxidation was catalyzed by FMOs in rat BMECs. Although FMO activity in rat BMECs was lower than that in SD rat normal hepatocytes (rtNHeps), we suggest that rat BMECs enzymes can convert substrates of exogenous origin for detoxification, indicating that BMECs are an important barrier for metabolic products besides hepatic cells.展开更多
Formate oxidase(FOx),which contains 8-formyl flavin adenine dinucleotide(FAD),exhibits a distinct advantage in utilizing ambient oxygen molecules for the oxidation of formic acid compared to other glucose-methanol-cho...Formate oxidase(FOx),which contains 8-formyl flavin adenine dinucleotide(FAD),exhibits a distinct advantage in utilizing ambient oxygen molecules for the oxidation of formic acid compared to other glucose-methanol-choline(GMC)oxidoreductase enzymes that contain only the standard FAD cofactor.The FOx-mediated conversion of FAD to 8-formyl FAD results in an approximate 10-fold increase in formate oxidase activity.However,the mechanistic details underlying the autocatalytic formation of 8-formyl FAD are still not well understood,which impedes further utilization of FOx.In this study,we employ molecular dynamics simulation,QM/MM umbrella sampling simulation,enzyme activity assay,site-directed mutagenesis,and spectroscopic analysis to elucidate the oxidation mechanism of FAD to 8-formyl FAD.Our results reveal that a catalytic water molecule,rather than any catalytic amino acids,serves as a general base to deprotonate the C8 methyl group on FAD,thus facilitating the formation of a quinone-methide tautomer intermediate.An oxygen molecule subsequently oxidizes this intermediate,resulting in a C8 methyl hydroperoxide anion that is protonated and dissociated to form OHC-RP and OH−.During the oxidation of FAD to 8-formyl FAD,the energy barrier for the rate-limiting step is calculated to be 22.8 kcal/mol,which corresponds to the required 14-hour transformation time observed experimentally.Further,the elucidated oxidation mechanism reveals that the autocatalytic formation of 8-formyl FAD depends on the proximal arginine and serine residues,R87 and S94,respectively.Enzymatic activity assay validates that the mutation of R87 to lysine reduces the kcat value to 75%of the wild-type,while the mutation to histidine results in a complete loss of activity.Similarly,the mutant S94I also leads to the deactivation of enzyme.This dependency arises because the nucleophilic OH−group and the quinone-methide tautomer intermediate are stabilized through the noncovalent interaction provided by R87 and S94.These findings not only explain the mechanistic details of each reaction step but also clarify the functional role of R87 and S94 during the oxidative maturation of 8-formyl FAD,thereby providing crucial theoretical support for the development of novel flavoenzymes with enhanced redox properties.展开更多
Methane monooxygenase(MMO)has attracted significant attention owing to its crucial role in the global carbon cycle;it impedes greenhouse effects by converting methane to methanol under ambient conditions.The water-sol...Methane monooxygenase(MMO)has attracted significant attention owing to its crucial role in the global carbon cycle;it impedes greenhouse effects by converting methane to methanol under ambient conditions.The water-soluble form of MMO(sMMO)has three essential components for the hydroxylation of methane:hydroxylase(MMOH),reductase(MMOR),and a regulatory(MMOB)component.MMOR consists of a flavin adenine dinucleotide(FAD)binding domain and a ferredoxin domain containing the[2Fe–2S]cluster for electron transfer from NADH to the di-iron sites in MMOH,which exerts its catalytic activity through O_(2)activation.Herein,the electronic structures of two cofactors,the FAD radical and[2Fe–2S]^(+),of reduced MMOR from Methylosinus sporium strain 5 were investigated.The results of multi-frequency and multi-technique electron paramagnetic resonance(EPR)spectroscopy of chemically reduced MMOR indicated the presence of a neutral flavin radical,and its detailed electronic structure was supported by density functional theory(DFT)calculations.The electronic and oxidation environments of[2Fe–2S]^(+)were further investigated using advanced EPR spectroscopy.Spectroscopic results confirmed that the oxidized state of Fe^(Ⅲ)is positioned near Cys50,which consists of a ferredoxin domain with a 2.7Ådistance between iron atoms.Our EPR spectroscopic results may provide a paradigm to elucidate the distribution of electronic densities of multiple cofactors in the enzyme,thus understanding its functional role.展开更多
Dear Editor,In hop(Humulus lupulus,Cannabaceae),the glandular trichomes of the female cones are highly efficient metabolic factories that accumulate specialized metabolites.These metabolites shape beer quality and mak...Dear Editor,In hop(Humulus lupulus,Cannabaceae),the glandular trichomes of the female cones are highly efficient metabolic factories that accumulate specialized metabolites.These metabolites shape beer quality and make hops indispensable to the brewing industry(Schonberger and Kostelecky,2011).While terpenoids define beer aroma,bitter acids(α-acids andβ-acids)confer the characteristic bitterness and were historically important for beer preservation.Among these,theα-acids-humulone,cohumulone,and adhumulone-are considered the‘‘soul’’of beer,serving as key ingredients that distinguish beer styles(Oladokun et al.,2017).Chemically,theα-acids undergo thermal isomerization during wort boiling to form iso-α-acids,the compounds responsible for beer bitterness(Figure 1A).展开更多
Objective To explore the effects of Xinglou Chengqi Decoction(XCD)on severe traumatic brain injury(sTBI)and its relationship with gut microbiota.Methods C57BL/6J mice were randomly allocated into sham,controlled corti...Objective To explore the effects of Xinglou Chengqi Decoction(XCD)on severe traumatic brain injury(sTBI)and its relationship with gut microbiota.Methods C57BL/6J mice were randomly allocated into sham,controlled cortical impact(CCI),and 3 doses of XCD(4.1,8.2,and 16.4 g/kg)groups by using a random number table,n=7 per group.A CCI device was employed to establish the TBI model.XCD was administered intragastrically for 3 consecutive days.The effects of XCD on post-sTBI neurological deficits and histopathology were assessed.The contribution of gut microbiota to XCD-mediated improvement in sTBI was investigated using antibiotic-treated TBI mice.The gut microbiota-dependent mechanisms of XCD in sTBI were explored through 16S rDNA sequencing and serum metabolomics.The mechanisms underlying the absorbed ingredients of XCD in sTBI were examined using network pharmacology and metabolomics.Finally,mice were divided into sham,CCI,and flavin adenine dinucleotide(FAD)-treated groups,n=10 per group.FAD was administered to sTBI mice via daily tail vein injection(830µg/kg)for 3 consecutive days to evaluate and verify its therapeutic effect.Results XCD significantly mitigated neurological impairments,neuronal damage,apoptosis,and blood-brain barrier disruption in CCI model mice(P<0.05 or P<0.01).The medium dose(8.2 g/kg)exhibited the greatest effect.The gut microbiota partly contributed to these protective effects.16S rDNA sequencing indicated that XCD promoted beneficial gut microbiota.Metabolomic analysis demonstrated that XCD regulated serum metabolic profiles,particularly FAD.Network pharmacology combined with metabolomics analysis revealed that the gut microbiota-independent components of XCD also targeted FAD in TBI.FAD exerted neuroprotective effects,improved energy metabolism,and promoted angiogenesis following TBI(P<0.05 or P<0.01).Conclusion XCD exerts neuroprotective effects on sTBI through both gut microbiota-dependent and-independent mechanisms,which highlight the therapeutic role of FAD.展开更多
Riboflavin （Rf） receptors bind and translocate Rf and its phosphorylated forms （e.g. flavin mononucleotide, FMN） into cells where they mediate various cellular metabolic pathways. Previously, we showed that FMN-co...Riboflavin （Rf） receptors bind and translocate Rf and its phosphorylated forms （e.g. flavin mononucleotide, FMN） into cells where they mediate various cellular metabolic pathways. Previously, we showed that FMN-coated ultrasmall superparamagnetic iron oxide （FLUSPIO） nanoparticles are suitable for labeling metabolically active cancer and endothelial cells in vitro. In this study, we focused on the in vivo application of FLUSPIO using prostate cancer xenografts. Size, charge, and chemical composition of FLUSPIO were evaluated. We explored the in vitro specificity of FLUSPIO for its cellular receptors using magnetic resonance imaging （MRI） and Prussian blue staining. Competitive binding experiments were performed in vivo by injecting free FMN in excess. Bio-distribution of FLUSPIO was determined by estimating iron content in organs and tumors using a colorimetric assay. AFM analysis and zeta potential measurements revealed a particulate morphology approximately 20-40 nm in size and a negative zeta potential （-24.23±0.15 mV） in water. X-ray photoelectron spectroscopy and time-of-flight secondary ion mass spectrometry data confirmed FMN present on the USPIO nanoparticle surface. FLUSPIO uptake in prostate cancer cells and human umbilical vein endothelial cells was significantly higher than that of control USPIO, while addition of excess of free FMN reduced accumulation. Similarly, in vivo MRI and histology showed specific FLUSPIO uptake by prostate cancer cells, tumor endothelial cells, and tumor-associated macrophages. Besides prominent tumor accumulation, FLUSPIO accumulated in the liver, spleen, lung, and skin. Hence, our data strengthen our hypothesis that targeting riboflavin receptors is an efficient approach to accumulate nanomedicines in tumors opening perspectives for the development of diagnostic and therapeutic systems.展开更多
Flavin-dependent halogenases (FDHs) are well known to introduce carbon halide bonds (mainly C–Cl and C–Br) into natural products with the assistance of a partner protein flavin reductase to generate reduced flavin (...Flavin-dependent halogenases (FDHs) are well known to introduce carbon halide bonds (mainly C–Cl and C–Br) into natural products with the assistance of a partner protein flavin reductase to generate reduced flavin (FADH_(2)or FMNH_(2)).Compared with the common chloride-and bromide-containing natural products (approximately 5,000 compounds),iodinated natural products(approximately 100 compounds) are very limited.Specific iodinases have also rarely been identified in nature to date.This study discovered a novel relationship between iodination and flavin reductases for the first time.Through mechanistic studies,it was identified that peroxide (H_(2)O_(2)) was released from the uncoupling reaction of flavin reductases and then reacted with iodide ions(I^(-)) to produce hypoiodous acid (IOH) for the final iodination.Furthermore,this study also unintentionally verified that the recently reported flavin-dependent iodinase Vir X1 from the marine virus and its two homologs (MBG and NCV) did not catalyze iodination in the in vitro biochemical system but likely belonged to a new phylogenetic clade in the tryptophan halogenase superfamily.As a consequence,actual flavin-dependent iodinases in nature remain to be discovered by the scientific community in the future.展开更多
Flavin-containing monooxygenase (FMO), like cytochrome P450 (CYP), is a monooxygenase that uses the reducing equivalents of NADPH to reduce one atom of molecular oxygen to water, while the other atom is used to oxidiz...Flavin-containing monooxygenase (FMO), like cytochrome P450 (CYP), is a monooxygenase that uses the reducing equivalents of NADPH to reduce one atom of molecular oxygen to water, while the other atom is used to oxidize the substrate. Recently, it was shown that some CYP isoforms have been subject to positive selection. However, it is unknown whether the highly conserved phase I detoxification enzyme, FMO, has undergone similar positive Darwinian selection. We used maximum-likelihood models of codon substitution, evolutionary fingerprinting, and cross species comparison to investigate the occurrence of adaptive evolution in FMO sequences. We used recent genomic data from a range of species, including vertebrates and invertebrates. We present the evidence for the occurrence of adaptive evolution in mammalian FMO 3, 4, 5, and fugu FMOs but not in mammalian FMO 1, FMO 2, frog FMOs, other fish FMOs and invertebrate FMOs. The sites under adaptive evolution were significantly associated with the insertion domain in mammalian FMO 5. We identified specific amino acid sites in FMOs 3–5 that are likely targets for selection based on the patterns of parallel amino acid change. The most likely role of adaptive evolution is the repair of mutations that permitted optimal NADP+ binding and improved catalytic efficiency. The occurrence of positive selection during the evolution of phase I detoxification enzymes such as FMOs 3–5 and fugu FMO suggests the occurrence of both high selection pressure acting on species within their unique habitats and significant changes in intensity and direction (forms of xenobiotics and drugs) resulting from changes in microhabitat and food.展开更多
基金Financial support for this research was provided by Modern Agro-industry Technology Research System of China(CARS-34)the Science and Technology Innovation Leading Talent in Central Plains(No.244200510010)the Outstanding Talents of Henan Agricultural University(No.30500636)。
文摘Background During the weaning phase,piglets are exposed to significant physiological and environmental stressors,which disrupt the balance of their intestinal microbiota and often lead to severe diarrhea.Previous studies have demonstrated that alfalfa fiber,derived from the stems and leaves of alfalfa,can effectively alleviate diarrhea in piglets.Additionally,multiple studies have highlighted the potential of fecal microbiota transplantation(FMT)in mitigating diarrhea in various models of intestinal diseases in young animals.However,the specific mechanisms by which FMT from targeted sources alleviates diarrhea in weaned piglets remain to be fully elucidated.Results In this study,FMT from donor piglets fed an alfalfa fiber-supplemented diet effectively alleviated diarrhea,improved intestinal morphology,and enhanced gut barrier function in weaned piglets.FMT further promoted the colonization of beneficial bacterial genera(including UCG-005,unclassified Lachnospiraceae,Lachnospiraceae AC2044 group,UCG-002,Candidatus Saccharimonas,and Lachnospiraceae ND3007 group)while inhibiting the detrimental genus Tyzzerella,consequently enhancing the production of short-chain fatty acids(SCFAs).Additionally,FMT upregulated riboflavin metabolism,leading to elevated flavin adenine dinucleotide(FAD)levels and increased glutathione reductase activity,thereby collectively attenuating lipopolysaccharide(LPS)-induced oxidative stress and contributing to intestinal health.Conclusions We found that FMT modulates the structure of the gut microbiota,enhances microbial diversity and composition,increases the production of SCFAs,and upregulates riboflavin metabolism to elevate FAD levels.These changes collectively enhance immune and antioxidant capacities,thereby alleviating diarrhea.
基金supported by the National Natural Science Foundation of China(Nos.52101078,U2006219)the National Key Research and Development Program of China(No.2020YFA0907300)+1 种基金the Fundamental Research Funds for the Central Universities of the Ministry of Education of China(Nos.N2102009,N2002019)Liaoning Revitalization Talents Program(No.XLYC1907158).
文摘Microbes can cause or accelerate metal corrosion,leading to huge losses in corrosion damages each year.Geobacter sulfurreducens is a representative electroactive bacterium in many soils,sediments,and wastew-ater systems.It has been confirmed to directly extract electrons from elemental metals.However,little is known about the effect of electron shuttles in G.sulfurreducens corrosion on stainless steel.In this study,we report that exogenous flavins promote iron-to-microbe electron transfer,accelerating micro-bial corrosion.G.sulfurreducens caused 1.3 times deeper pits and increased electron uptake(with 2 times increase of i_(corr))from stainless steel when riboflavin was added to the culture medium.OmcS-deficient mutant data suggest that G.sulfurreducens utilizes riboflavin as a bound-cofactor in outer membrane c-type cytochromes.The finding that,in the presence of microbes,riboflavin can substantially accelerate corrosion highlights the role of flavin redox cycling for enhanced iron-to-microbe electron transfer by G.sulfurreducens and provides new insights in microbial corrosion.
文摘The aim of this study was to examine whether flavin-containing monooxygenase (FMO) protein was expressed in cultured rat brain microvascular endothelial cells (BMECs), which constitute the blood-brain barrier (BBB), and whether N-oxide from the tertiary amine, d-chlorpheniramine, was formed by FMO in rat BMECs. BMECs were isolated and cultured from the brains of three-week-old male Wistar rats. The expression of FMO1, FMO2 and FMO5 proteins was confirmed in rat BMECs by western blotting analysis using polyclonal anti-FMO antibodies, but FMO3 and FMO4 proteins were not found in the rat BBB. Moreover, N-oxide of d-chlorpheniramine was formed in rat BMECs. The intrinsic clearance value for N-oxidation at pH 8.4 was higher than that at pH 7.4. Inhibition of N-oxide formation by methimazole was found to be the best model of competitive inhibition yielding an apparent Ki value of 0.53 μmol/L, suggesting that N-oxidation was catalyzed by FMOs in rat BMECs. Although FMO activity in rat BMECs was lower than that in SD rat normal hepatocytes (rtNHeps), we suggest that rat BMECs enzymes can convert substrates of exogenous origin for detoxification, indicating that BMECs are an important barrier for metabolic products besides hepatic cells.
基金supported by the National Natural Science Foundation of China(32201030,32271319 and 32071267)the Science and Technology Department of Jilin Province(20230402041GH and YDZJ202301ZYTS537)+2 种基金the Education Department of Jilin Province(JJKH20220970KJ)the Development and Reform Commission of Jilin Province(2023C015)the Fundamental Research Funds of the Central Universities in China(2024-JCXK-11).
文摘Formate oxidase(FOx),which contains 8-formyl flavin adenine dinucleotide(FAD),exhibits a distinct advantage in utilizing ambient oxygen molecules for the oxidation of formic acid compared to other glucose-methanol-choline(GMC)oxidoreductase enzymes that contain only the standard FAD cofactor.The FOx-mediated conversion of FAD to 8-formyl FAD results in an approximate 10-fold increase in formate oxidase activity.However,the mechanistic details underlying the autocatalytic formation of 8-formyl FAD are still not well understood,which impedes further utilization of FOx.In this study,we employ molecular dynamics simulation,QM/MM umbrella sampling simulation,enzyme activity assay,site-directed mutagenesis,and spectroscopic analysis to elucidate the oxidation mechanism of FAD to 8-formyl FAD.Our results reveal that a catalytic water molecule,rather than any catalytic amino acids,serves as a general base to deprotonate the C8 methyl group on FAD,thus facilitating the formation of a quinone-methide tautomer intermediate.An oxygen molecule subsequently oxidizes this intermediate,resulting in a C8 methyl hydroperoxide anion that is protonated and dissociated to form OHC-RP and OH−.During the oxidation of FAD to 8-formyl FAD,the energy barrier for the rate-limiting step is calculated to be 22.8 kcal/mol,which corresponds to the required 14-hour transformation time observed experimentally.Further,the elucidated oxidation mechanism reveals that the autocatalytic formation of 8-formyl FAD depends on the proximal arginine and serine residues,R87 and S94,respectively.Enzymatic activity assay validates that the mutation of R87 to lysine reduces the kcat value to 75%of the wild-type,while the mutation to histidine results in a complete loss of activity.Similarly,the mutant S94I also leads to the deactivation of enzyme.This dependency arises because the nucleophilic OH−group and the quinone-methide tautomer intermediate are stabilized through the noncovalent interaction provided by R87 and S94.These findings not only explain the mechanistic details of each reaction step but also clarify the functional role of R87 and S94 during the oxidative maturation of 8-formyl FAD,thereby providing crucial theoretical support for the development of novel flavoenzymes with enhanced redox properties.
基金supported by the National Research Foundation of Korea(NRF-2017M3D1A1039380 and NRF-2017R1A2B4008691)to S.H.K.S.J.Lthe Ministry of Education(2017R1A6A1A03015876)the C1 Gas Refinery Program(NRF-2015M3D3D3A1A01064876)for funding.
文摘Methane monooxygenase(MMO)has attracted significant attention owing to its crucial role in the global carbon cycle;it impedes greenhouse effects by converting methane to methanol under ambient conditions.The water-soluble form of MMO(sMMO)has three essential components for the hydroxylation of methane:hydroxylase(MMOH),reductase(MMOR),and a regulatory(MMOB)component.MMOR consists of a flavin adenine dinucleotide(FAD)binding domain and a ferredoxin domain containing the[2Fe–2S]cluster for electron transfer from NADH to the di-iron sites in MMOH,which exerts its catalytic activity through O_(2)activation.Herein,the electronic structures of two cofactors,the FAD radical and[2Fe–2S]^(+),of reduced MMOR from Methylosinus sporium strain 5 were investigated.The results of multi-frequency and multi-technique electron paramagnetic resonance(EPR)spectroscopy of chemically reduced MMOR indicated the presence of a neutral flavin radical,and its detailed electronic structure was supported by density functional theory(DFT)calculations.The electronic and oxidation environments of[2Fe–2S]^(+)were further investigated using advanced EPR spectroscopy.Spectroscopic results confirmed that the oxidized state of Fe^(Ⅲ)is positioned near Cys50,which consists of a ferredoxin domain with a 2.7Ådistance between iron atoms.Our EPR spectroscopic results may provide a paradigm to elucidate the distribution of electronic densities of multiple cofactors in the enzyme,thus understanding its functional role.
基金supported by the National Natural Science Foundation of China(grant no.32470265 to G.W.)the China Postdoctoral Science Foundation(grant no.2021M693388 to C.F.)We sincerely thank Dr.Jonathan E.Page(University of British Columbia,Canada)and Dr.Shawn M.Clark(National Research Council of Canada)for their valuable and insightful discussions on the hops project.
文摘Dear Editor,In hop(Humulus lupulus,Cannabaceae),the glandular trichomes of the female cones are highly efficient metabolic factories that accumulate specialized metabolites.These metabolites shape beer quality and make hops indispensable to the brewing industry(Schonberger and Kostelecky,2011).While terpenoids define beer aroma,bitter acids(α-acids andβ-acids)confer the characteristic bitterness and were historically important for beer preservation.Among these,theα-acids-humulone,cohumulone,and adhumulone-are considered the‘‘soul’’of beer,serving as key ingredients that distinguish beer styles(Oladokun et al.,2017).Chemically,theα-acids undergo thermal isomerization during wort boiling to form iso-α-acids,the compounds responsible for beer bitterness(Figure 1A).
基金Supported by the Leadership Training Project of Hunan Traditional Chinese Medicine(No.2022-04)Fundamental Research Funds for the Central Universities of Central South University(Nos.2023ZZTS0564 and 2024ZZTS0928)。
文摘Objective To explore the effects of Xinglou Chengqi Decoction(XCD)on severe traumatic brain injury(sTBI)and its relationship with gut microbiota.Methods C57BL/6J mice were randomly allocated into sham,controlled cortical impact(CCI),and 3 doses of XCD(4.1,8.2,and 16.4 g/kg)groups by using a random number table,n=7 per group.A CCI device was employed to establish the TBI model.XCD was administered intragastrically for 3 consecutive days.The effects of XCD on post-sTBI neurological deficits and histopathology were assessed.The contribution of gut microbiota to XCD-mediated improvement in sTBI was investigated using antibiotic-treated TBI mice.The gut microbiota-dependent mechanisms of XCD in sTBI were explored through 16S rDNA sequencing and serum metabolomics.The mechanisms underlying the absorbed ingredients of XCD in sTBI were examined using network pharmacology and metabolomics.Finally,mice were divided into sham,CCI,and flavin adenine dinucleotide(FAD)-treated groups,n=10 per group.FAD was administered to sTBI mice via daily tail vein injection(830µg/kg)for 3 consecutive days to evaluate and verify its therapeutic effect.Results XCD significantly mitigated neurological impairments,neuronal damage,apoptosis,and blood-brain barrier disruption in CCI model mice(P<0.05 or P<0.01).The medium dose(8.2 g/kg)exhibited the greatest effect.The gut microbiota partly contributed to these protective effects.16S rDNA sequencing indicated that XCD promoted beneficial gut microbiota.Metabolomic analysis demonstrated that XCD regulated serum metabolic profiles,particularly FAD.Network pharmacology combined with metabolomics analysis revealed that the gut microbiota-independent components of XCD also targeted FAD in TBI.FAD exerted neuroprotective effects,improved energy metabolism,and promoted angiogenesis following TBI(P<0.05 or P<0.01).Conclusion XCD exerts neuroprotective effects on sTBI through both gut microbiota-dependent and-independent mechanisms,which highlight the therapeutic role of FAD.
文摘Riboflavin （Rf） receptors bind and translocate Rf and its phosphorylated forms （e.g. flavin mononucleotide, FMN） into cells where they mediate various cellular metabolic pathways. Previously, we showed that FMN-coated ultrasmall superparamagnetic iron oxide （FLUSPIO） nanoparticles are suitable for labeling metabolically active cancer and endothelial cells in vitro. In this study, we focused on the in vivo application of FLUSPIO using prostate cancer xenografts. Size, charge, and chemical composition of FLUSPIO were evaluated. We explored the in vitro specificity of FLUSPIO for its cellular receptors using magnetic resonance imaging （MRI） and Prussian blue staining. Competitive binding experiments were performed in vivo by injecting free FMN in excess. Bio-distribution of FLUSPIO was determined by estimating iron content in organs and tumors using a colorimetric assay. AFM analysis and zeta potential measurements revealed a particulate morphology approximately 20-40 nm in size and a negative zeta potential （-24.23±0.15 mV） in water. X-ray photoelectron spectroscopy and time-of-flight secondary ion mass spectrometry data confirmed FMN present on the USPIO nanoparticle surface. FLUSPIO uptake in prostate cancer cells and human umbilical vein endothelial cells was significantly higher than that of control USPIO, while addition of excess of free FMN reduced accumulation. Similarly, in vivo MRI and histology showed specific FLUSPIO uptake by prostate cancer cells, tumor endothelial cells, and tumor-associated macrophages. Besides prominent tumor accumulation, FLUSPIO accumulated in the liver, spleen, lung, and skin. Hence, our data strengthen our hypothesis that targeting riboflavin receptors is an efficient approach to accumulate nanomedicines in tumors opening perspectives for the development of diagnostic and therapeutic systems.
基金the National Natural Science Foundation of China(21632007,21661140002 for S.Lin81903525 for Y.Zhang)+1 种基金Research Fund for High-level Talents of Xinxiang Medical University(300-505272)Open Funding Project of State Key Laboratory of Microbial Metabolism,Shanghai Jiao Tong University(MMLKF2011)。
文摘Flavin-dependent halogenases (FDHs) are well known to introduce carbon halide bonds (mainly C–Cl and C–Br) into natural products with the assistance of a partner protein flavin reductase to generate reduced flavin (FADH_(2)or FMNH_(2)).Compared with the common chloride-and bromide-containing natural products (approximately 5,000 compounds),iodinated natural products(approximately 100 compounds) are very limited.Specific iodinases have also rarely been identified in nature to date.This study discovered a novel relationship between iodination and flavin reductases for the first time.Through mechanistic studies,it was identified that peroxide (H_(2)O_(2)) was released from the uncoupling reaction of flavin reductases and then reacted with iodide ions(I^(-)) to produce hypoiodous acid (IOH) for the final iodination.Furthermore,this study also unintentionally verified that the recently reported flavin-dependent iodinase Vir X1 from the marine virus and its two homologs (MBG and NCV) did not catalyze iodination in the in vitro biochemical system but likely belonged to a new phylogenetic clade in the tryptophan halogenase superfamily.As a consequence,actual flavin-dependent iodinases in nature remain to be discovered by the scientific community in the future.
基金supported by the National Science and Technology Major Project of the Ministry of Science and Technology of China (2008ZX10005-004)Educational Commission of Liaoning Province of China (2009A120)China Postdoctoral Science Foundation (20080440019 and 200902069)
文摘Flavin-containing monooxygenase (FMO), like cytochrome P450 (CYP), is a monooxygenase that uses the reducing equivalents of NADPH to reduce one atom of molecular oxygen to water, while the other atom is used to oxidize the substrate. Recently, it was shown that some CYP isoforms have been subject to positive selection. However, it is unknown whether the highly conserved phase I detoxification enzyme, FMO, has undergone similar positive Darwinian selection. We used maximum-likelihood models of codon substitution, evolutionary fingerprinting, and cross species comparison to investigate the occurrence of adaptive evolution in FMO sequences. We used recent genomic data from a range of species, including vertebrates and invertebrates. We present the evidence for the occurrence of adaptive evolution in mammalian FMO 3, 4, 5, and fugu FMOs but not in mammalian FMO 1, FMO 2, frog FMOs, other fish FMOs and invertebrate FMOs. The sites under adaptive evolution were significantly associated with the insertion domain in mammalian FMO 5. We identified specific amino acid sites in FMOs 3–5 that are likely targets for selection based on the patterns of parallel amino acid change. The most likely role of adaptive evolution is the repair of mutations that permitted optimal NADP+ binding and improved catalytic efficiency. The occurrence of positive selection during the evolution of phase I detoxification enzymes such as FMOs 3–5 and fugu FMO suggests the occurrence of both high selection pressure acting on species within their unique habitats and significant changes in intensity and direction (forms of xenobiotics and drugs) resulting from changes in microhabitat and food.
