Rice sheath blight(RSB)is a major destructive disease impeding rice production.Identifying key germplasm resources with increased resistance remains a challenge.However,the mechanisms underlying disease resistance are...Rice sheath blight(RSB)is a major destructive disease impeding rice production.Identifying key germplasm resources with increased resistance remains a challenge.However,the mechanisms underlying disease resistance are not yet fully understood.Cytochrome P450 monooxygenases(CYP450s)serve biosynthesis and metabolic detoxification functions in plants,but there is limited information about their role in the response induced by RSB.This study demonstrated that CYT02 belongs to the CYP73A100 subfamily and is a typical member of the CYP450s.Overexpression(OE)in rice of the cytochrome P450 monooxygenase cyt02 conferred increased resistance to RSB and increased vegetative tillering.Cyt02 may increase RSB resistance by regulating plant hormone synthesis,regulate reactive oxygen species(ROS)by coordinating the activity of antioxidant enzymes,and initiate phytoalexin synthesis in response to fungal infection.These research findings have laid a foundation for a deeper understanding of the function of cyt02 and offered a potential target gene for breeding rice varieties resistant to sheath blight.展开更多
Methane(CH_(4))has a higher heat capacity(104.9 kcal/mol)than carbon dioxide(CO_(2)),and this has inspired research aimed at reducing methane levels to retard global warming.Hydroxylation under ambient conditions thro...Methane(CH_(4))has a higher heat capacity(104.9 kcal/mol)than carbon dioxide(CO_(2)),and this has inspired research aimed at reducing methane levels to retard global warming.Hydroxylation under ambient conditions through methanotrophs can provide crucial information for understanding the harsh C-H activation of methane.Soluble methane monooxygenase(sMMO)belongs to the bacterial multi-component monooxygenase superfamily and requires hydroxylase(MMOH),regulatory(MMOB),and reductase(MMOR)components.Recent structural and biophysical studies have demonstrated that these components accelerate and retard methane hydroxylation in MMOH through protein-protein interactions.Complex structures of sMMO,including MMOH-MMOB and MMOH-MMOD,illustrate how these regulatory and inhibitory components orchestrate the di-iron active sites located within the four-helix bundles of MMOH,specifically at the docking surface known as the canyon region.In addition,recent biophysical studies have demonstrated the role of MmoR,aσ54-dependent transcriptional regulator,in regulating sMMO expression.This perspective article introduces remarkable discoveries in recent reports on sMMO components that are crucial for understanding sMMO expression and activities.Our findings provide insight into how sMMO components interact with MMOH to control methane hydroxylation,shedding light on the mechanisms governing sMMO expression and the interactions between activating enzymes and promoters.展开更多
Betaine is a very effective osmoprotectant found in many organisms. In high plants, betaine is synthesized by oxidation of choline in two sequential steps: choline-->betaine aldehyde-->betaine. The first step is...Betaine is a very effective osmoprotectant found in many organisms. In high plants, betaine is synthesized by oxidation of choline in two sequential steps: choline-->betaine aldehyde-->betaine. The first step is catalyzed by choline monooxygenase (CMO). In this study, the full-length CMO cDNA (1 820 bp) was cloned from halophyte Suaeda liaotungensis Kitag by RT-PCR and RACE. It included a 123 bp 5' UTR, a 368 bp 3' UTR and a 1 329 bp open reading frame encoding a 442-amino-acid polypeptide with 77%, 72% and 74% sequence identity compared to CMOs from spinach, sugar beet and Atriplex hortensis, respectively. The CMO open reading frame (ORF) was cloned and the plant expression vector pBI121-CMO was constructed. It was transferred into tobacco ( Nicotiana tabacum L. ev. 89) via Agrobacterium mediation. PCR and Southern blotting analysis showed that the CMO gene was integrated into tobacco genome. Transgenic tobacco plants contained higher amount of betaine than that of control plants and were able to survive on MS medium containing 250 mmol/L NaCl. Relative electronic conductivity demonstrated less membrane damage in transgenic plants as in the wild type.展开更多
Plants synthesize the osmoprotectant glycine betaine (GB) via choline→betaine aldehyde→glycine be- taine[1]. Two enzymes are involved in the pathway choline monooxygenase (CMO) and betaine aldehyde dehydrogenase (BA...Plants synthesize the osmoprotectant glycine betaine (GB) via choline→betaine aldehyde→glycine be- taine[1]. Two enzymes are involved in the pathway choline monooxygenase (CMO) and betaine aldehyde dehydrogenase (BADH). A full length CMO cDNA (1,643bp) was cloned from Amaranthus tricolor. The open reading frame encoded a 442-amino acid polypeptide, which showed 69% identity with CMOs in Spina- cia oleracea L. and Beta vulgaris L. DNA gel blot analysis indicated the presence of one copy of CMO gene in the A. tricolor genome. The expressions of CMO and BADH proteins in A.tricolor leaves significantly increased under salinization, drought and heat stress (42℃), as determined by immunoblot analysis, but did not respond to cold stress (4℃), or exogenous ABA application. The increase of GB content in leaves was parallel to CMO and BADH contents.展开更多
The coupled effects of nitrogen source and methane monooxygenase（MMO） on the growth and poly-β-hydroxybutyrate（PHB） accumulation capacity of methanotrophs were explored.The ammonia-supplied methanotrophs expressi...The coupled effects of nitrogen source and methane monooxygenase（MMO） on the growth and poly-β-hydroxybutyrate（PHB） accumulation capacity of methanotrophs were explored.The ammonia-supplied methanotrophs expressing soluble MMO（s MMO） grew at the highest rate, while N2-fixing bacteria expressing particulate MMO（p MMO） grew at the lowest rate. Further study showed that more hydroxylamine and nitrite was formed by ammonia-supplied bacteria containing p MMO, which might cause their slightly lower growth rate. The highest PHB content（51.0%） was obtained under nitrogen-limiting conditions with the inoculation of nitrate-supplied bacteria containing p MMO. Ammoniasupplied bacteria also accumulated a higher content of PHB（45.2%） with the expression of p MMO, while N2-fixing bacteria containing p MMO only showed low PHB production capacity（32.1%）. The maximal PHB contents of bacteria expressing s MMO were low, with no significant change under different nitrogen source conditions. The low MMO activity,low cell growth rate and low PHB production capacity of methanotrophs continuously cultivated with N2 with the expression of p MMO were greatly improved in the cyclic NO3-N2 cultivation regime, indicating that long-term deficiency of nitrogen sources was detrimental to the activity of methanotrophs expressing pMMO.展开更多
[ Objective] This study aimed to investigate the function of aliD gene in the inactivation of AHLs. [ Method ] A bacterial isolate, Acinetobacter sp. CA2 from soil, is capable of inactivation of AHLs. A gene designed ...[ Objective] This study aimed to investigate the function of aliD gene in the inactivation of AHLs. [ Method ] A bacterial isolate, Acinetobacter sp. CA2 from soil, is capable of inactivation of AHLs. A gene designed as aliD, which is responsible for AHL-quenching activity and exhibits high similarity with Mo- nooxygenase genes, was cloned from the genomic library of Acinetobacter sp. CA2. [ Result ] The aliD gene in-frame deletion mutant, CA2 AliD, impaired its AHLs inactivating function when mixed with N-（3-oxooctanoyl） -L-homosefine lactone （30C8-HSL）. Expression of AliD in plant pathogenic bacterium Pectobacterium ca- rotovorum subsp, carotovorum Z3-3 significantly reduced the AHLs production and the extracellular pectolytic enzyme activities, and attenuated soft rot disease symptoms on the plants tested, including potato, Chinese cabbage, radish and cabbage. [ Conclusion ] Our study suggests that the aliD gene complemented strain CA2-AliD showes a similar AHLs inactivating function.展开更多
Based on part of a known cDNA sequence of Suaeda Liaotungensis choline monooxygenase, the authors successfully cloned the 5′ cDNA end of Suaeda Lianotungensis choline monooxygenase using Inverse PCR RACE with a speci...Based on part of a known cDNA sequence of Suaeda Liaotungensis choline monooxygenase, the authors successfully cloned the 5′ cDNA end of Suaeda Lianotungensis choline monooxygenase using Inverse PCR RACE with a specially designed 5′-phosphated RT primer and two pairs of specific inverse PCR primers. Compared with the anchored PCR RACE, inverse PCR RACE has better specificity and higher amplification.展开更多
A transformation procedure of choline monooxygenase(CMO) gene, involved in stress tolerance, was established in white pine embryogenic tissue by using A. tumefaciens C58/pMP90. The CMO cDNA fragment(1.3 kb) was genera...A transformation procedure of choline monooxygenase(CMO) gene, involved in stress tolerance, was established in white pine embryogenic tissue by using A. tumefaciens C58/pMP90. The CMO cDNA fragment(1.3 kb) was generated by Reverse Transcription-Polymerase Chain Reaction (RT-PCR) with primers based on the report sequence of CMO in gene bank. A chimerical gene composed of the cauliflower mosaic virus (CaMV) 35S promoter fused to CMO cDNA and β-glucuronidase (GUS-marker gene) was transferred into Ti-derived disarmed binary vector pBI121. The new vector, p35SCMOp, was transferred into Agrobacterium tumefaciens C58/pMP90 by freeze-thaw method. Somatic embryogenesis (SE) initiation of Pinus. Strobus L. and Pinus.Koraiensis Sieb. et Zucc. depended on the manipulation of plant growth regulator (PGR) concentrations in the GLH culture medium. Transgenic embryos and regenerated plants of two Pine species were produced after co-culture of embryogenic tissue with the disarmed strain of A. tumefaciens C58/pMP90/ p35SCMOp and selected on medium containing 25mg/L kanamycin. The transformed embryogenic tissue was initially confirmed by histochemical GUS assay followed by PCR. One copy of T-DNA was detected by transgenic lines analysis in Pinus. Strobus L. and transgenic plants were regenerated for two species using modified protocols for maturation and germination of somatic embryos.展开更多
The effects on ethoxyresorufin O-deethylase （EROD）, NADPH-cytochrome c reductase and NADH cytochrorne b5 reductase activities of digestive gland in Meretrix meretrix exposed to tributyltin （TBT） at environmental l...The effects on ethoxyresorufin O-deethylase （EROD）, NADPH-cytochrome c reductase and NADH cytochrorne b5 reductase activities of digestive gland in Meretrix meretrix exposed to tributyltin （TBT） at environmental levels （0.1,1.0,10.0 ng/dm as stannum concentration）,in experimental condition, were evaluated. The EROD, NADH cytochrorne b5 reductase activities were significantly inhibited after exposure to 10.0 ng/dm^3 TBT for 8 and 20 d, the NADPH cytochrorne c reductase activities were significantly inhibited after exposure to 0.1,1.0 and 10.0 ng/dm TBT for 8 d and to 1.0 and 10.0 ng/dm for 20 d, as compared with the matched control, while NADH cytochrorne b5 reductases and NADPH cytochrome c reductase activities were induced after exposure to 10.0 ng/dm^3 TBT for 2 d. The EROD activity in the 10 ng/dm^3 group,and the NADH cytochrome b5 reductases activities in 1.0 and 10.0 ng/dm groups, were significantly induced when transferred to clean recovery tanks for 7 d. The three enzymic activities in the clams exposed to TBT were recovered to the level corresponding to that of the control group after transfer to clean recovery tanks for 20 d. NADPH cytochrome c reductase activity in Meretrix meretrix seems to be more sensitive to exposure of TBT than that of the EROD and NADH cytochrome b5 reductases. The results suggest that induction and inhibition by TBT to the monooxygenase system enzymic activity in Meretrix meretrix would simultaneously exist. The enzymic activities were inhibited by exposure for a long time. The results suggest that inhibition of the monooxygenase system should be an indication of the exposure to environmentally relevant concentrations of TBT for a long time.展开更多
A methane monooxygenase-like dinuclear complex of iron (Ⅲ) with a chiral heptadentate ligand has been synthesed and tested for catalytic ability for asymmetric epoxidation of styrene with iodosylbenzene. The results ...A methane monooxygenase-like dinuclear complex of iron (Ⅲ) with a chiral heptadentate ligand has been synthesed and tested for catalytic ability for asymmetric epoxidation of styrene with iodosylbenzene. The results show that the complex exhibits many qualitative features, such as electronic spectrum. catalytic behavior. and asymmetric induction similar to those of methane monooxygenase and catalyzes styrene epoxidation to R- (+)-styrene oxide in 9. 3% yield and 72. 9% optical yield.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
Atkanesulfonate monooxygenase SsuD facilitates the desulfonation reaction of alkane sulfonates to release sulfite and corresponding aldehydes/ketones. Oxygen is activated by the reduced flavin. One oxygen atom is to f...Atkanesulfonate monooxygenase SsuD facilitates the desulfonation reaction of alkane sulfonates to release sulfite and corresponding aldehydes/ketones. Oxygen is activated by the reduced flavin. One oxygen atom is to fi＇om water and the other oxygen atom is to from aldehydes/ketones. The oxidized flavin is regenerated after water is formed. The chemical biomimetic system was established according to the preliminary mechanism of alkanesulfonate monooxygenase and the cyclic mechanism was proposed for the formation ofaldehydes/ketones. Two oxygen atoms from the reduced flavin to form C（4a）-peroxy-flavin. The oxygen atom connected with C（4a） abstracts one electron from the neighbouring oxygen to transfer one oxygen atom to C1 ofalkanesulfonates and abstracts one hydrogen from C1 ofalkanesulfonates to break C 1-H bond. Hydroxy-flavin was produced by the above cyclic mechanism. Alkansulfonate monooxygenase SsuD does not directly involve in the reaction. It only supplies some comfortable environment to facilitate the target reactiorL展开更多
Regioselective and enantioselective hydroxylation of propargylic C-H bonds are useful reactions but often lack appropriate catalysts.Here a green and efficient asymmetric hydroxylation of primary and secondary C-H bon...Regioselective and enantioselective hydroxylation of propargylic C-H bonds are useful reactions but often lack appropriate catalysts.Here a green and efficient asymmetric hydroxylation of primary and secondary C-H bonds at propargylic positions has been established.A series of optically active propargylic alcohols were prepared with high regio-and enantioselectivity(up to 99%ee)under mild reaction conditions by using P450tol,while the C≡C bonds in the molecule remained unreacted.This protocol provides a green and practical method for constructing enantiomerically chiral propargylic alcohols.In addition,we also demonstrated that the biohydroxylation strategy was able to scaled up to 2.25 mmol scale with the production of chiral propargyl alcohol 2a at a yield of 196 mg with 96%ee,which’s an important synthetic intermediate of antifungal drug Ravuconazole.展开更多
Dear Editor,Glyceollins are isoflavonoid-derived metabolites produced by soybean that hold great promise in improving human and animal health.The recurrent use of antibiotics in animal production has led to the emerge...Dear Editor,Glyceollins are isoflavonoid-derived metabolites produced by soybean that hold great promise in improving human and animal health.The recurrent use of antibiotics in animal production has led to the emergence of antibiotic resistance,which is causing a global challenge for public health and food systems.Owing to their broad-spectrum antimicrobial properties,glyceollins are being proposed to replace antibiotics in the swine industry to reduce antibiotic-resistant microorganisms in the food supply(Zavadil,2020;Ika Irianti et al.,2023).展开更多
Transmembrane alkane monooxygenase(AlkB)-type monooxygenases,especially AlkB2 monooxygenases,are crucial for aerobic degradation of the medium-to-long-chain n-alkanes in hydrocarbon-utilizing microorganisms.In this st...Transmembrane alkane monooxygenase(AlkB)-type monooxygenases,especially AlkB2 monooxygenases,are crucial for aerobic degradation of the medium-to-long-chain n-alkanes in hydrocarbon-utilizing microorganisms.In this study,we identified a GntR/VanR transcription regulator AlkR of Pseudomonas aeruginosa SJTD-1 involved in the negative regulation of AlkB2 and deciphered its nature of DNA binding and ligand release.The deletion of alkR enhanced the transcription levels of the alkB2 gene and the utilization efficiency of the medium-to-long-chain n-alkanes by strain SJTD-1.The dimer of AlkR recognizes and binds to a conserved palindromic motif in the promoter of the alkB2 gene,and structural symmetry is vital for DNA binding and transcription repression.The long-chain fatty acyl coenzyme A compounds can release AlkR and stimulate transcription of alkB2,reflecting the effect of alkane catabolic metabolites.Structural insights unveiled that the arginine residues and scaffold residues of AlkR are critical for DNA binding.Further bioinformatics analysis of AlkR revealed the widespread VanR–AlkB couples distributed in Pseudomonadaceae with high conservation in the sequences of functional genes and intergenic regions,highlighting a conserved regulatory pattern for n-alkane utilization across this family.These findings demonstrate the regulatory mechanism and structural basis of GntR/VanR transcription regulators in modulating n-alkane biodegradation and provide valuable insights in improving the bioremediation efficiency of hydrocarbon pollution.展开更多
Background:Focal segmental glomerulosclerosis (FSGS)is a kidney disease that is commonly associated with proteinuria and the progressive loss of renal function,which is characterized by podocyte injury and the depleti...Background:Focal segmental glomerulosclerosis (FSGS)is a kidney disease that is commonly associated with proteinuria and the progressive loss of renal function,which is characterized by podocyte injury and the depletion and collapse of glomerular capillary segments.The pathogenesis of FSGS has not been completely elucidated;however,recent advances in molecular genetics have provided increasing evidence that podocyte structural and functional disruption is central to FSGS pathogenesis.Here,we identified a patient with FSGS and aimed to characterize the pathogenic gene and verify its mechanism. Methods:Using next-generation sequencing and Sanger sequencing,we screened the causative gene that was linked to FSGS in this study.The patient's total blood RNA was extracted to validate the messenger RNA (mRNA)expression of coenzyme Q10 monooxygenase 6(COQ6)and validated it by immunohistochemistry.COQ6 knockdown in podocytes was performed in vitro with small interfering RNA, and then,F-actin was determined using immunofluorescence staining.Cell apoptosis was evaluated by flow cytometry,the expression of active caspase-3was determined by Western blot,and mitochondrial function was detected by MitoSOX. Results:Using whole-exome sequencing and Sanger sequencing,we screened a new causative gene,COQ6,NM_182480:exonl:c.G41A: p.W14X.The mRNA expression of COQ6 in the proband showed decreased.Moreover,the expression of COQ6,which was validated by immunohistochemistry,also had the same change in the proband.Finally,we focused on the COQ6 gene to clarify the mechanism of podocyte injury.Flow cytometry showed significantly increased in apoptotic podocytes,and Western blotting showed increases in active caspase-3in si-COQ6 podocytes.Meanwhile,reactive oxygen species (ROS)levels were increased and F-actin immunofluorescence was irregularly distributed in the si-COQ6 group. Conclusions:This study reported a possible mechanism for FSGS and suggested that a new mutation in COQ6,which could cause respiratory chain defect,increase the generation of ROS,destroy the podocyte cytoskeleton,and induce apoptosis.It provides basic theoretical basis for the screening of FSGS in the future.展开更多
Lytic polysaccharide monooxygenases(LPMOs)are important enzymes that boost the hydrolysis of recalcitrant polysaccharides,such as chitin.They are found extensively in different insect species and are classified as aux...Lytic polysaccharide monooxygenases(LPMOs)are important enzymes that boost the hydrolysis of recalcitrant polysaccharides,such as chitin.They are found extensively in different insect species and are classified as auxiliary activities family 15(AA15)LPMOs(LPMO15).Some of them were identified from the insect midgut and proven to act on chitin.However,knowledge about their physiological roles during insect growth and development remains limited.Here,we found that midgut-specific LPMO15s are widely distributed in different insect orders,such as the orthopteran Locusta migratoria and the lepidopteran Bombyx mori.Using L.migratoria as a model insect,the function of midgut-specific LmLPMO15-3 during development was investigated.Double-stranded RNA-mediated downregulation of LmLPMO15-3 expression at the 4th or 5th instar nymph stage severely decreased the survival rate and resulted in lethal phenotypes.Hematoxylin and eosin staining results indicated that the deficient individuals exhibited incompletely digested peritrophic matrix(PM),which suggested that LmLPMO15-3 is essential for the deconstruction of the PM during molting.This study provides direct evidence of the physiological importance of a midgut-specific LPMO15 during insect development.As L.migratoria is one of the most destructive agricultural pests,LmLPMO15-3 is a potential target for pest management.展开更多
Roxithromycin, livestock growth promoter, as widely used medicine and arouses concern because its occurrence and persistence in soil environments. However, effects of roxithromycin in higher plants are still vague. Ac...Roxithromycin, livestock growth promoter, as widely used medicine and arouses concern because its occurrence and persistence in soil environments. However, effects of roxithromycin in higher plants are still vague. Accordingly, we hypothesized that roxithromycin-con- taminated soil may exhibits ecotoxicological effects in wheat （Triticum aestivum）. In this study, effects induced by a gradient concentration of roxithromycin stress （0.01, 0.1, 1, 10, and 100 mg.kg-1） was investigated in a 7-d soil test in T. aestivum. Results indicated that the specific activity of cytochrome P450 （CYP450） monooxygenase was decreased dramatically with the concentration of roxithro- mycin in soil. The IC50 value was 8.78mg.kg-1 of roxithromycin. On the contrary, the growth related endpoints （i.e., the germination percentage, the biomass and the height）, the content related endpoints （i.e., soluble protein content and CYP450 content）, and the superoxide dismutase （SOD） activity failed to reveal the roxithromy- cin-induced effects. Further analysis revealed that the CYP450 monooxygenase specific activity reduction was enzymatic mechanism mediated, other than oxidative stress induced. We conclude that the soil roxithromycin declined the CYP450 monooxygenase activity in T. aestivum by the inhibition of the enzymatic mechanism. Further efforts can include, but are not limited to, investigation of joint effects induced by combined exposure of roxithromycin and the pesticides and evalua- tion of the similar effects in other higher plants.展开更多
Glycyrrhiza uralensis is frequently used in traditional Chinese medicine. This plant contains a large amount of effective constituents, including triterpenoids and flavonoids. Among them, glycyrrhizin is believed to b...Glycyrrhiza uralensis is frequently used in traditional Chinese medicine. This plant contains a large amount of effective constituents, including triterpenoids and flavonoids. Among them, glycyrrhizin is believed to be the marker compound to evaluate the quality of G. uralensis based on Chinese Pharmacopoeia. Many studies showed that glycyrrhizin possesses various pharmacological activities, such as antibacterial, antiviral, antitumor, anti-inflammatory, and immune-stimulating activities. In this paper, we summarized the cloning, characterization, expression, and polymorphism analysis of several functional genes involved in glycyrrhizin biosynthesis in G. uralensis.展开更多
基金supported by the Sichuan Province International Science and Technology Innovation Cooperation(2024YFHZ0299)the Project of Science and Technology Department of Sichuan Province(2022YFH0031)Chengdu Science and Technology Bureau(2024-YF05-02168-SN).
文摘Rice sheath blight(RSB)is a major destructive disease impeding rice production.Identifying key germplasm resources with increased resistance remains a challenge.However,the mechanisms underlying disease resistance are not yet fully understood.Cytochrome P450 monooxygenases(CYP450s)serve biosynthesis and metabolic detoxification functions in plants,but there is limited information about their role in the response induced by RSB.This study demonstrated that CYT02 belongs to the CYP73A100 subfamily and is a typical member of the CYP450s.Overexpression(OE)in rice of the cytochrome P450 monooxygenase cyt02 conferred increased resistance to RSB and increased vegetative tillering.Cyt02 may increase RSB resistance by regulating plant hormone synthesis,regulate reactive oxygen species(ROS)by coordinating the activity of antioxidant enzymes,and initiate phytoalexin synthesis in response to fungal infection.These research findings have laid a foundation for a deeper understanding of the function of cyt02 and offered a potential target gene for breeding rice varieties resistant to sheath blight.
基金This research was supported by"Regional Innovation Strategy"(2023RIS-008)and"C1 Gas Refinery Program"(NRF-2015M3D3D3A1A01064876)through the National Research Foundation of Koreafunded by the Ministry of Education(NRF-2017R1A6A1A03015876).
文摘Methane(CH_(4))has a higher heat capacity(104.9 kcal/mol)than carbon dioxide(CO_(2)),and this has inspired research aimed at reducing methane levels to retard global warming.Hydroxylation under ambient conditions through methanotrophs can provide crucial information for understanding the harsh C-H activation of methane.Soluble methane monooxygenase(sMMO)belongs to the bacterial multi-component monooxygenase superfamily and requires hydroxylase(MMOH),regulatory(MMOB),and reductase(MMOR)components.Recent structural and biophysical studies have demonstrated that these components accelerate and retard methane hydroxylation in MMOH through protein-protein interactions.Complex structures of sMMO,including MMOH-MMOB and MMOH-MMOD,illustrate how these regulatory and inhibitory components orchestrate the di-iron active sites located within the four-helix bundles of MMOH,specifically at the docking surface known as the canyon region.In addition,recent biophysical studies have demonstrated the role of MmoR,aσ54-dependent transcriptional regulator,in regulating sMMO expression.This perspective article introduces remarkable discoveries in recent reports on sMMO components that are crucial for understanding sMMO expression and activities.Our findings provide insight into how sMMO components interact with MMOH to control methane hydroxylation,shedding light on the mechanisms governing sMMO expression and the interactions between activating enzymes and promoters.
文摘Betaine is a very effective osmoprotectant found in many organisms. In high plants, betaine is synthesized by oxidation of choline in two sequential steps: choline-->betaine aldehyde-->betaine. The first step is catalyzed by choline monooxygenase (CMO). In this study, the full-length CMO cDNA (1 820 bp) was cloned from halophyte Suaeda liaotungensis Kitag by RT-PCR and RACE. It included a 123 bp 5' UTR, a 368 bp 3' UTR and a 1 329 bp open reading frame encoding a 442-amino-acid polypeptide with 77%, 72% and 74% sequence identity compared to CMOs from spinach, sugar beet and Atriplex hortensis, respectively. The CMO open reading frame (ORF) was cloned and the plant expression vector pBI121-CMO was constructed. It was transferred into tobacco ( Nicotiana tabacum L. ev. 89) via Agrobacterium mediation. PCR and Southern blotting analysis showed that the CMO gene was integrated into tobacco genome. Transgenic tobacco plants contained higher amount of betaine than that of control plants and were able to survive on MS medium containing 250 mmol/L NaCl. Relative electronic conductivity demonstrated less membrane damage in transgenic plants as in the wild type.
文摘Plants synthesize the osmoprotectant glycine betaine (GB) via choline→betaine aldehyde→glycine be- taine[1]. Two enzymes are involved in the pathway choline monooxygenase (CMO) and betaine aldehyde dehydrogenase (BADH). A full length CMO cDNA (1,643bp) was cloned from Amaranthus tricolor. The open reading frame encoded a 442-amino acid polypeptide, which showed 69% identity with CMOs in Spina- cia oleracea L. and Beta vulgaris L. DNA gel blot analysis indicated the presence of one copy of CMO gene in the A. tricolor genome. The expressions of CMO and BADH proteins in A.tricolor leaves significantly increased under salinization, drought and heat stress (42℃), as determined by immunoblot analysis, but did not respond to cold stress (4℃), or exogenous ABA application. The increase of GB content in leaves was parallel to CMO and BADH contents.
基金supported by the National Key Scientific and Technology Project for Water Pollution Treatment of China(No.2012ZX07202006)the National Natural Science Foundation of China(No.21477014)
文摘The coupled effects of nitrogen source and methane monooxygenase（MMO） on the growth and poly-β-hydroxybutyrate（PHB） accumulation capacity of methanotrophs were explored.The ammonia-supplied methanotrophs expressing soluble MMO（s MMO） grew at the highest rate, while N2-fixing bacteria expressing particulate MMO（p MMO） grew at the lowest rate. Further study showed that more hydroxylamine and nitrite was formed by ammonia-supplied bacteria containing p MMO, which might cause their slightly lower growth rate. The highest PHB content（51.0%） was obtained under nitrogen-limiting conditions with the inoculation of nitrate-supplied bacteria containing p MMO. Ammoniasupplied bacteria also accumulated a higher content of PHB（45.2%） with the expression of p MMO, while N2-fixing bacteria containing p MMO only showed low PHB production capacity（32.1%）. The maximal PHB contents of bacteria expressing s MMO were low, with no significant change under different nitrogen source conditions. The low MMO activity,low cell growth rate and low PHB production capacity of methanotrophs continuously cultivated with N2 with the expression of p MMO were greatly improved in the cyclic NO3-N2 cultivation regime, indicating that long-term deficiency of nitrogen sources was detrimental to the activity of methanotrophs expressing pMMO.
基金Supported by the Natural Science Foundation of China(30871666,31071725)Special Fund for Agro-scientific Research in the Public Interest(201003066)the National High Technology Research and Development Program of China("863" Program)(2011AA10A205)
文摘[ Objective] This study aimed to investigate the function of aliD gene in the inactivation of AHLs. [ Method ] A bacterial isolate, Acinetobacter sp. CA2 from soil, is capable of inactivation of AHLs. A gene designed as aliD, which is responsible for AHL-quenching activity and exhibits high similarity with Mo- nooxygenase genes, was cloned from the genomic library of Acinetobacter sp. CA2. [ Result ] The aliD gene in-frame deletion mutant, CA2 AliD, impaired its AHLs inactivating function when mixed with N-（3-oxooctanoyl） -L-homosefine lactone （30C8-HSL）. Expression of AliD in plant pathogenic bacterium Pectobacterium ca- rotovorum subsp, carotovorum Z3-3 significantly reduced the AHLs production and the extracellular pectolytic enzyme activities, and attenuated soft rot disease symptoms on the plants tested, including potato, Chinese cabbage, radish and cabbage. [ Conclusion ] Our study suggests that the aliD gene complemented strain CA2-AliD showes a similar AHLs inactivating function.
文摘Based on part of a known cDNA sequence of Suaeda Liaotungensis choline monooxygenase, the authors successfully cloned the 5′ cDNA end of Suaeda Lianotungensis choline monooxygenase using Inverse PCR RACE with a specially designed 5′-phosphated RT primer and two pairs of specific inverse PCR primers. Compared with the anchored PCR RACE, inverse PCR RACE has better specificity and higher amplification.
文摘A transformation procedure of choline monooxygenase(CMO) gene, involved in stress tolerance, was established in white pine embryogenic tissue by using A. tumefaciens C58/pMP90. The CMO cDNA fragment(1.3 kb) was generated by Reverse Transcription-Polymerase Chain Reaction (RT-PCR) with primers based on the report sequence of CMO in gene bank. A chimerical gene composed of the cauliflower mosaic virus (CaMV) 35S promoter fused to CMO cDNA and β-glucuronidase (GUS-marker gene) was transferred into Ti-derived disarmed binary vector pBI121. The new vector, p35SCMOp, was transferred into Agrobacterium tumefaciens C58/pMP90 by freeze-thaw method. Somatic embryogenesis (SE) initiation of Pinus. Strobus L. and Pinus.Koraiensis Sieb. et Zucc. depended on the manipulation of plant growth regulator (PGR) concentrations in the GLH culture medium. Transgenic embryos and regenerated plants of two Pine species were produced after co-culture of embryogenic tissue with the disarmed strain of A. tumefaciens C58/pMP90/ p35SCMOp and selected on medium containing 25mg/L kanamycin. The transformed embryogenic tissue was initially confirmed by histochemical GUS assay followed by PCR. One copy of T-DNA was detected by transgenic lines analysis in Pinus. Strobus L. and transgenic plants were regenerated for two species using modified protocols for maturation and germination of somatic embryos.
文摘The effects on ethoxyresorufin O-deethylase （EROD）, NADPH-cytochrome c reductase and NADH cytochrorne b5 reductase activities of digestive gland in Meretrix meretrix exposed to tributyltin （TBT） at environmental levels （0.1,1.0,10.0 ng/dm as stannum concentration）,in experimental condition, were evaluated. The EROD, NADH cytochrorne b5 reductase activities were significantly inhibited after exposure to 10.0 ng/dm^3 TBT for 8 and 20 d, the NADPH cytochrorne c reductase activities were significantly inhibited after exposure to 0.1,1.0 and 10.0 ng/dm TBT for 8 d and to 1.0 and 10.0 ng/dm for 20 d, as compared with the matched control, while NADH cytochrorne b5 reductases and NADPH cytochrome c reductase activities were induced after exposure to 10.0 ng/dm^3 TBT for 2 d. The EROD activity in the 10 ng/dm^3 group,and the NADH cytochrome b5 reductases activities in 1.0 and 10.0 ng/dm groups, were significantly induced when transferred to clean recovery tanks for 7 d. The three enzymic activities in the clams exposed to TBT were recovered to the level corresponding to that of the control group after transfer to clean recovery tanks for 20 d. NADPH cytochrome c reductase activity in Meretrix meretrix seems to be more sensitive to exposure of TBT than that of the EROD and NADH cytochrome b5 reductases. The results suggest that induction and inhibition by TBT to the monooxygenase system enzymic activity in Meretrix meretrix would simultaneously exist. The enzymic activities were inhibited by exposure for a long time. The results suggest that inhibition of the monooxygenase system should be an indication of the exposure to environmentally relevant concentrations of TBT for a long time.
文摘A methane monooxygenase-like dinuclear complex of iron (Ⅲ) with a chiral heptadentate ligand has been synthesed and tested for catalytic ability for asymmetric epoxidation of styrene with iodosylbenzene. The results show that the complex exhibits many qualitative features, such as electronic spectrum. catalytic behavior. and asymmetric induction similar to those of methane monooxygenase and catalyzes styrene epoxidation to R- (+)-styrene oxide in 9. 3% yield and 72. 9% optical yield.
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.
文摘Atkanesulfonate monooxygenase SsuD facilitates the desulfonation reaction of alkane sulfonates to release sulfite and corresponding aldehydes/ketones. Oxygen is activated by the reduced flavin. One oxygen atom is to fi＇om water and the other oxygen atom is to from aldehydes/ketones. The oxidized flavin is regenerated after water is formed. The chemical biomimetic system was established according to the preliminary mechanism of alkanesulfonate monooxygenase and the cyclic mechanism was proposed for the formation ofaldehydes/ketones. Two oxygen atoms from the reduced flavin to form C（4a）-peroxy-flavin. The oxygen atom connected with C（4a） abstracts one electron from the neighbouring oxygen to transfer one oxygen atom to C1 ofalkanesulfonates and abstracts one hydrogen from C1 ofalkanesulfonates to break C 1-H bond. Hydroxy-flavin was produced by the above cyclic mechanism. Alkansulfonate monooxygenase SsuD does not directly involve in the reaction. It only supplies some comfortable environment to facilitate the target reactiorL
基金financial support from the National Natural Science Foundation of China(No.32271537 and 22061049)the Science and Technology Department of Guizhou province(QKHJCZK2021-036 and QKHRCPTGCC-2023-003)+1 种基金the Science and Technology Department of Zunyi(ZSKRPT-2020-5,ZSKH-2018-3,ZSKRPT-2021-5)Zunyi Medical University(QKH-2018-5772-014).
文摘Regioselective and enantioselective hydroxylation of propargylic C-H bonds are useful reactions but often lack appropriate catalysts.Here a green and efficient asymmetric hydroxylation of primary and secondary C-H bonds at propargylic positions has been established.A series of optically active propargylic alcohols were prepared with high regio-and enantioselectivity(up to 99%ee)under mild reaction conditions by using P450tol,while the C≡C bonds in the molecule remained unreacted.This protocol provides a green and practical method for constructing enantiomerically chiral propargylic alcohols.In addition,we also demonstrated that the biohydroxylation strategy was able to scaled up to 2.25 mmol scale with the production of chiral propargyl alcohol 2a at a yield of 196 mg with 96%ee,which’s an important synthetic intermediate of antifungal drug Ravuconazole.
基金supported by Agriculture and Agri-Food Canada’s Abase(J-002364)the ASC-09 Soybean Cluster Activity#7A(J-002080)grants to S.D.
文摘Dear Editor,Glyceollins are isoflavonoid-derived metabolites produced by soybean that hold great promise in improving human and animal health.The recurrent use of antibiotics in animal production has led to the emergence of antibiotic resistance,which is causing a global challenge for public health and food systems.Owing to their broad-spectrum antimicrobial properties,glyceollins are being proposed to replace antibiotics in the swine industry to reduce antibiotic-resistant microorganisms in the food supply(Zavadil,2020;Ika Irianti et al.,2023).
基金supported by the National Key Research and Development Program of China(2021YFA0909500)the National Natural Science Foundation of China(32370107 and 32170106)the Natural Science Foundation of Shanghai(23ZR1431300).
文摘Transmembrane alkane monooxygenase(AlkB)-type monooxygenases,especially AlkB2 monooxygenases,are crucial for aerobic degradation of the medium-to-long-chain n-alkanes in hydrocarbon-utilizing microorganisms.In this study,we identified a GntR/VanR transcription regulator AlkR of Pseudomonas aeruginosa SJTD-1 involved in the negative regulation of AlkB2 and deciphered its nature of DNA binding and ligand release.The deletion of alkR enhanced the transcription levels of the alkB2 gene and the utilization efficiency of the medium-to-long-chain n-alkanes by strain SJTD-1.The dimer of AlkR recognizes and binds to a conserved palindromic motif in the promoter of the alkB2 gene,and structural symmetry is vital for DNA binding and transcription repression.The long-chain fatty acyl coenzyme A compounds can release AlkR and stimulate transcription of alkB2,reflecting the effect of alkane catabolic metabolites.Structural insights unveiled that the arginine residues and scaffold residues of AlkR are critical for DNA binding.Further bioinformatics analysis of AlkR revealed the widespread VanR–AlkB couples distributed in Pseudomonadaceae with high conservation in the sequences of functional genes and intergenic regions,highlighting a conserved regulatory pattern for n-alkane utilization across this family.These findings demonstrate the regulatory mechanism and structural basis of GntR/VanR transcription regulators in modulating n-alkane biodegradation and provide valuable insights in improving the bioremediation efficiency of hydrocarbon pollution.
文摘Background:Focal segmental glomerulosclerosis (FSGS)is a kidney disease that is commonly associated with proteinuria and the progressive loss of renal function,which is characterized by podocyte injury and the depletion and collapse of glomerular capillary segments.The pathogenesis of FSGS has not been completely elucidated;however,recent advances in molecular genetics have provided increasing evidence that podocyte structural and functional disruption is central to FSGS pathogenesis.Here,we identified a patient with FSGS and aimed to characterize the pathogenic gene and verify its mechanism. Methods:Using next-generation sequencing and Sanger sequencing,we screened the causative gene that was linked to FSGS in this study.The patient's total blood RNA was extracted to validate the messenger RNA (mRNA)expression of coenzyme Q10 monooxygenase 6(COQ6)and validated it by immunohistochemistry.COQ6 knockdown in podocytes was performed in vitro with small interfering RNA, and then,F-actin was determined using immunofluorescence staining.Cell apoptosis was evaluated by flow cytometry,the expression of active caspase-3was determined by Western blot,and mitochondrial function was detected by MitoSOX. Results:Using whole-exome sequencing and Sanger sequencing,we screened a new causative gene,COQ6,NM_182480:exonl:c.G41A: p.W14X.The mRNA expression of COQ6 in the proband showed decreased.Moreover,the expression of COQ6,which was validated by immunohistochemistry,also had the same change in the proband.Finally,we focused on the COQ6 gene to clarify the mechanism of podocyte injury.Flow cytometry showed significantly increased in apoptotic podocytes,and Western blotting showed increases in active caspase-3in si-COQ6 podocytes.Meanwhile,reactive oxygen species (ROS)levels were increased and F-actin immunofluorescence was irregularly distributed in the si-COQ6 group. Conclusions:This study reported a possible mechanism for FSGS and suggested that a new mutation in COQ6,which could cause respiratory chain defect,increase the generation of ROS,destroy the podocyte cytoskeleton,and induce apoptosis.It provides basic theoretical basis for the screening of FSGS in the future.
基金supported by the National Natural Science Foundation of China(31872972,31830076)the Shenzhen Science and Technology Program(KQTD20180411143628272).
文摘Lytic polysaccharide monooxygenases(LPMOs)are important enzymes that boost the hydrolysis of recalcitrant polysaccharides,such as chitin.They are found extensively in different insect species and are classified as auxiliary activities family 15(AA15)LPMOs(LPMO15).Some of them were identified from the insect midgut and proven to act on chitin.However,knowledge about their physiological roles during insect growth and development remains limited.Here,we found that midgut-specific LPMO15s are widely distributed in different insect orders,such as the orthopteran Locusta migratoria and the lepidopteran Bombyx mori.Using L.migratoria as a model insect,the function of midgut-specific LmLPMO15-3 during development was investigated.Double-stranded RNA-mediated downregulation of LmLPMO15-3 expression at the 4th or 5th instar nymph stage severely decreased the survival rate and resulted in lethal phenotypes.Hematoxylin and eosin staining results indicated that the deficient individuals exhibited incompletely digested peritrophic matrix(PM),which suggested that LmLPMO15-3 is essential for the deconstruction of the PM during molting.This study provides direct evidence of the physiological importance of a midgut-specific LPMO15 during insect development.As L.migratoria is one of the most destructive agricultural pests,LmLPMO15-3 is a potential target for pest management.
基金This research is financially supported by the National Natural Science Foundation of China as a general project （Grant No. 31170473） and a joint Guangdong project （No. U1133006）, and by the Ministry of Education of China as an innovative team project （No. IRT 13024）.
文摘Roxithromycin, livestock growth promoter, as widely used medicine and arouses concern because its occurrence and persistence in soil environments. However, effects of roxithromycin in higher plants are still vague. Accordingly, we hypothesized that roxithromycin-con- taminated soil may exhibits ecotoxicological effects in wheat （Triticum aestivum）. In this study, effects induced by a gradient concentration of roxithromycin stress （0.01, 0.1, 1, 10, and 100 mg.kg-1） was investigated in a 7-d soil test in T. aestivum. Results indicated that the specific activity of cytochrome P450 （CYP450） monooxygenase was decreased dramatically with the concentration of roxithro- mycin in soil. The IC50 value was 8.78mg.kg-1 of roxithromycin. On the contrary, the growth related endpoints （i.e., the germination percentage, the biomass and the height）, the content related endpoints （i.e., soluble protein content and CYP450 content）, and the superoxide dismutase （SOD） activity failed to reveal the roxithromy- cin-induced effects. Further analysis revealed that the CYP450 monooxygenase specific activity reduction was enzymatic mechanism mediated, other than oxidative stress induced. We conclude that the soil roxithromycin declined the CYP450 monooxygenase activity in T. aestivum by the inhibition of the enzymatic mechanism. Further efforts can include, but are not limited to, investigation of joint effects induced by combined exposure of roxithromycin and the pesticides and evalua- tion of the similar effects in other higher plants.
基金Beijing Project for Young Talents(YETP0819)National Science Foundation of China(81503181)
文摘Glycyrrhiza uralensis is frequently used in traditional Chinese medicine. This plant contains a large amount of effective constituents, including triterpenoids and flavonoids. Among them, glycyrrhizin is believed to be the marker compound to evaluate the quality of G. uralensis based on Chinese Pharmacopoeia. Many studies showed that glycyrrhizin possesses various pharmacological activities, such as antibacterial, antiviral, antitumor, anti-inflammatory, and immune-stimulating activities. In this paper, we summarized the cloning, characterization, expression, and polymorphism analysis of several functional genes involved in glycyrrhizin biosynthesis in G. uralensis.
