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1.
Small ; 20(7): e2303502, 2024 Feb.
Article in English | MEDLINE | ID: mdl-37840447

ABSTRACT

Borophene, a promising material with potential applications in electronics, energy storage, and sensors, is successfully grown as a monolayer on Ag(111), Cu(111), and Au(111) surfaces using molecular beam epitaxy. The growth of two-dimensional borophene on Ag(111) and Au(111) is proposed to occur via surface adsorption and boron segregation, respectively. However, the growth mode of borophene on Cu(111) remains unclear. To elucidate this, scanning tunneling microscopy in conjunction with theoretical calculations is used to study the phase transformation of boron nanostructures under post-annealing treatments. Results show that by elevating the substrate temperature, boron nanostructures undergo an evolution from amorphous boron to striped-phase borophene (η = 1/6) adhering to the Cu ⟨ 1 1 ¯ 0 ⟩ $\langle {1\bar{1}0} \rangle $ step edge, and finally to irregularly shaped ß-type borophene (η = 5/36) either on the substrate surface or embedded in the topmost Cu layer. dI/dV spectra recorded near the borophene/Cu lateral interfaces indicate that the striped-phase borophene is a metastable phase, requiring more buckling and electron transfer to stabilize the crystal structure. These findings offer not only an in-depth comprehension of the ß-type borophene formation on Cu(111), but also hold potential for enabling borophene synthesis on weakly-binding semiconducting or insulating substrates with 1D active defects.

2.
J Sci Food Agric ; 104(10): 5999-6007, 2024 Aug 15.
Article in English | MEDLINE | ID: mdl-38436580

ABSTRACT

BACKGROUND: Isomaltulose is a 'generally recognized as safe' ingredient and is widely used in the food, pharmaceutical and chemical industries. The exploration and development of efficient technologies is essential for enhancing isomaltulose yield. RESULTS: In the present study, a simple and efficient surface display platform mediated by a non-yeast signal peptide was developed in Yarrowia lipolytica and utilized to efficiently produce isomaltulose from sucrose. We discovered that the signal peptide SP1 of sucrose isomerase from Pantoea dispersa UQ68J (PdSI) could guide SIs anchoring to the cell surface of Y. lipolytica, demonstrating a novel and simple cell surface display strategy. Furthermore, the PdSI expression level was significantly increased through optimizing the promoters and multi-site integrating genes into chromosome. The final strain gained 451.7 g L-1 isomaltulose with a conversion rate of 90.3% and a space-time yield of 50.2 g L-1 h-1. CONCLUSION: The present study provides an efficient way for manufacturing isomaltulose with a high space-time yield. This heterogenous signal peptide-mediated cell surface display strategy featured with small fusion tag (approximately 2.2 kDa of SP1), absence of enzyme leakage in fermentation broth and ample room for optimization, providing a convenient way to construct whole-cell biocatalysts to synthesize other products and broadening the array of molecular toolboxes accessible for engineering Y. lipolytica. © 2024 Society of Chemical Industry.


Subject(s)
Isomaltose , Protein Sorting Signals , Yarrowia , Yarrowia/genetics , Yarrowia/metabolism , Isomaltose/metabolism , Isomaltose/analogs & derivatives , Metabolic Engineering , Pantoea/genetics , Pantoea/metabolism , Pantoea/enzymology , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Fungal Proteins/genetics , Fungal Proteins/metabolism , Cell Surface Display Techniques , Glucosyltransferases/genetics , Glucosyltransferases/metabolism
3.
Molecules ; 28(1)2022 Dec 26.
Article in English | MEDLINE | ID: mdl-36615394

ABSTRACT

The advent of graphene opens up the research into two-dimensional (2D) materials, which are considered revolutionary materials. Due to its unique geometric structure, graphene exhibits a series of exotic physical and chemical properties. In addition, single-element-based 2D materials (Xenes) have garnered tremendous interest. At present, 16 kinds of Xenes (silicene, borophene, germanene, phosphorene, tellurene, etc.) have been explored, mainly distributed in the third, fourth, fifth, and sixth main groups. The current methods to prepare monolayers or few-layer 2D materials include epitaxy growth, mechanical exfoliation, and liquid phase exfoliation. Although two Xenes (aluminene and indiene) have not been synthesized due to the limitations of synthetic methods and the stability of Xenes, other Xenes have been successfully created via elaborate artificial design and synthesis. Focusing on elemental 2D materials, this review mainly summarizes the recently reported work about tuning the electronic, optical, mechanical, and chemical properties of Xenes via surface modifications, achieved using controllable approaches (doping, adsorption, strain, intercalation, phase transition, etc.) to broaden their applications in various fields, including spintronics, electronics, optoelectronics, superconducting, photovoltaics, sensors, catalysis, and biomedicines. These advances in the surface modification of Xenes have laid a theoretical and experimental foundation for the development of 2D materials and their practical applications in diverse fields.

4.
Biochem Biophys Res Commun ; 579: 54-61, 2021 11 19.
Article in English | MEDLINE | ID: mdl-34587555

ABSTRACT

1,2-ß-Mannobiose phosphorylases (1,2-ß-MBPs) from glycoside hydrolase 130 (GH130) family are important bio-catalysts in glycochemistry applications owing to their ability in synthesizing oligomannans. Here, we report the crystal structure of a thermostable 1,2-ß-MBP from Thermoanaerobacter sp. X-514 termed Teth514_1789 to reveal the molecular basis of its higher thermostability and mechanism of action. We also solved the enzyme complexes of mannose, mannose-1-phosphate (M1P) and 1,4-ß-mannobiose to manifest the enzyme-substrate interaction networks of three main subsites. Notably, a Zn ion that should be derived from crystallization buffer was found in the active site and coordinates the phosphate moiety of M1P. Nonetheless, this Zn-coordination should reflect an inhibitory status as supplementing Zn severely impairs the enzyme activity. These results indicate that the effects of metal ions should be taken into consideration when applying Teth514_1789 and other related enzymes. Based on the structure, a reliable model of Teth514_1788 that shares 61.7% sequence identity to Teth514_1789 but displays a different substrate preference was built. Analyzing the structural features of these two closely related enzymes, we hypothesized that the length of a loop fragment that covers the entrance of the catalytic center might regulate the substrate selectivity. In conclusion, these information provide in-depth understanding of GH130 1,2-ß-MBPs and should serve as an important guidance for enzyme engineering for further applications.


Subject(s)
Thermoanaerobacter/enzymology , beta-Mannosidase/chemistry , Binding Sites , Catalysis , Catalytic Domain , Glycoside Hydrolases/chemistry , Ions , Ligands , Mannans/chemistry , Mannose/chemistry , Mannosephosphates/chemistry , Phosphorylases/chemistry , Plasmids/metabolism , Protein Conformation , Reproducibility of Results , Static Electricity , Temperature , Zinc/chemistry
5.
Genome ; 64(1): 51-61, 2021 Jan.
Article in English | MEDLINE | ID: mdl-33105087

ABSTRACT

Clostridium butyricum is an anaerobic bacterium that inhabits broad niches. Clostridium butyricum is known for its production of butyrate, 1,3-propanediol, and hydrogen. This study aimed to present a comparative pangenome analysis of 24 strains isolated from different niches. We sequenced and annotated the genome of C. butyricum 3-3 isolated from the Chinese baijiu ecosystem. The pangenome of C. butyricum was open. The core genome, accessory genome, and strain-specific genes comprised 1011, 4543, and 1473 genes, respectively. In the core genome, Carbohydrate metabolism was the largest category, and genes in the biosynthetic pathway of butyrate and glycerol metabolism were conserved (in the core or soft-core genome). Furthermore, the 1,3-propanediol operon existed in 20 strains. In the accessory genome, numerous mobile genetic elements belonging to the Replication, recombination, and repair (L) category were identified. In addition, genome islands were identified in all 24 strains, ranging from 2 (strain KNU-L09) to 53 (strain SU1), and phage sequences were found in 17 of the 24 strains. This study provides an important genomic framework that could pave the way for the exploration of C. butyricum and future studies on the genetic diversification of C. butyricum.


Subject(s)
Clostridium butyricum/genetics , Clostridium butyricum/metabolism , Butyrates/metabolism , Carbohydrate Metabolism , Clostridium butyricum/classification , Fermentation , Genes, Bacterial/genetics , Genome Size , Metabolic Networks and Pathways/genetics , Operon , Phylogeny , Propylene Glycols , Sequence Analysis, DNA
6.
Metab Eng ; 61: 215-224, 2020 09.
Article in English | MEDLINE | ID: mdl-32623008

ABSTRACT

Starch/cellulose has become the major feedstock for manufacturing biofuels and biochemicals because of their abundance and sustainability. In this study, we presented an artificially designed "starch-mannose-fermentation" biotransformation process through coupling the advantages of in vivo and in vitro metabolic engineering strategies together. Starch was initially converted into mannose via an in vitro metabolic engineering biosystem, and then mannose was fermented by engineered microorganisms for biomanufacturing valuable mannosyl compounds. The in vitro metabolic engineering biosystem based on phosphorylation/dephosphorylation reactions was thermodynamically favorable and the conversion rate reached 81%. The mannose production using whole-cell biocatalysts reached 75.4 g/L in a 30-L reactor, indicating the potential industrial application. Furthermore, the produced mannose in the reactor was directly served as feedstock for the fermentation process to bottom-up produced 19.2 g/L mannosyl-oligosaccharides (MOS) and 7.2 g/L mannosylglycerate (MG) using recombinant Corynebacterium glutamicum strains. Notably, such a mannose fermentation process facilitated the synthesis of MOS, which has not been achieved under glucose fermentation and improved MG production by 2.6-fold than that using the same C-mole of glucose. This approach also allowed access to produce other kinds of mannosyl derivatives from starch.


Subject(s)
Bioreactors , Corynebacterium glutamicum , Glyceric Acids , Mannose/analogs & derivatives , Metabolic Engineering , Microorganisms, Genetically-Modified , Starch/metabolism , Corynebacterium glutamicum/genetics , Corynebacterium glutamicum/growth & development , Mannose/biosynthesis , Mannose/genetics , Microorganisms, Genetically-Modified/genetics , Microorganisms, Genetically-Modified/growth & development
7.
Appl Environ Microbiol ; 85(2)2019 01 15.
Article in English | MEDLINE | ID: mdl-30389762

ABSTRACT

Herein, we present a biocatalytic method to produce raffinose and stachyose using sucrose as the substrate. An in vitro multienzyme system was developed using five enzymes, namely, sucrose synthase (SUS), UDP-glucose 4-epimerase (GalE), galactinol synthase (GS), raffinose synthase (RS), and stachyose synthase (STS), and two intermedia, namely, UDP and inositol, which can be recycled. This reaction system produced 11.1 mM raffinose using purified enzymes under optimal reaction conditions and substrate concentrations. Thereafter, a stepwise cascade reaction strategy was employed to circumvent the instability of RS and STS in this system, and a 4.2-fold increase in raffinose production was observed. The enzymatic cascade reactions were then conducted using cell extracts to avoid the need for enzyme purification and supplementation with UDP. Such modification further increased raffinose production to 86.6 mM and enabled the synthesis of 61.1 mM stachyose. The UDP turnover number reached 337. Finally, inositol in the reaction system was recycled five times, and 255.8 mM raffinose (128.9 g/liter) was obtained.IMPORTANCE Soybean oligosaccharides (SBOS) have elicited considerable attention because of their potential applications in the pharmaceutical, cosmetics, and food industries. This study demonstrates an alternative method to produce raffinose and stachyose, which are the major bioactive components of SBOS, from sucrose via an in vitro enzyme system. High concentrations of galactinol, raffinose, and stachyose were synthesized with the aid of a stepwise cascade reaction process, which can successfully address the issue of mismatched enzyme characteristics of an in vitro metabolic engineering platform. The biocatalytic approach presented in this work may enable the synthesis of other valuable galactosyl oligosaccharides, such as verbascose and higher homologs, which are difficult to obtain through plant extraction.


Subject(s)
Bacterial Proteins/metabolism , Multienzyme Complexes/metabolism , Oligosaccharides/biosynthesis , Plant Proteins/metabolism , Raffinose/biosynthesis , Sucrose/metabolism , Arabidopsis/enzymology , Escherichia coli/enzymology
8.
Biotechnol Bioeng ; 116(4): 745-756, 2019 04.
Article in English | MEDLINE | ID: mdl-30597517

ABSTRACT

D-Allulose 3-epimerase (DAE) has been applied to produce D-allulose, a low-calorie and functional sweetener. In this study, a new DAE from Paenibacillus senegalensis was characterized in Escherichia coli. Furthermore, we presented a tandem isoenzyme gene expression strategy to express multiple DAEs in one cell and construct food-grade expression systems based on Corynebacterium glutamicum. Seventeen expression cassettes based on three DAE genes from different organisms were constructed. Among all recombinant strains, DAE16 harboring three DAE genes in an expression vector exhibited the highest enzyme activity with 22.7 U/mg. Whole-cell transformation of DAE16 produced 225 g/L D-allulose with a volumetric productivity of 353 g·g -1 ·hr -1 . The catalytic efficiency of strain C-DAE9 integrating total 11 DAE genes in chromosome was 16.4-fold higher than strains carrying one DAE. Fed-batch culture of C-DAE9 gave enzyme activity of 44,700 U/L. We also expressed a thermostable invertase in C. glutamicum and obtained enzyme activity of 29 U/mg. Immobilized cells expressing DAE or invertase exhibited 80% of retained activity after 30 cycles of catalytic reactions. Those immobilized cells were coupled to produce 61.2 g/L D-allulose from cane molasses in a two-step reaction process. This study provided an efficient approach for enzyme preparation and allowed access to produce D-allulose from other abundant and low-cost feedstock enriched with sucrose.


Subject(s)
Bacterial Proteins/genetics , Corynebacterium glutamicum/genetics , Escherichia coli/genetics , Fructose/metabolism , Paenibacillus/genetics , Racemases and Epimerases/genetics , Bacterial Proteins/metabolism , Corynebacterium glutamicum/metabolism , Escherichia coli/metabolism , Fructose/analysis , Gene Expression , Genes, Bacterial , Industrial Microbiology , Isoenzymes/metabolism , Molasses/analysis , Paenibacillus/metabolism , Phylogeny , Racemases and Epimerases/metabolism , Saccharum/chemistry , Saccharum/metabolism , Transformation, Genetic
9.
Environ Res ; 166: 167-174, 2018 10.
Article in English | MEDLINE | ID: mdl-29886393

ABSTRACT

A TiO2/MWCNTs/Al2O3/NF (TiO2 nanoparticles/multi-walled carbon nanotubes /Al2O3-modified nickel foams) composite filter was prepared through grown of MWCNTs upon Al2O3-modified nickel foams using chemical vapor deposition (CVD) method and then coated TiO2 nanoparticles onto MWCNTs using dip-coating method. The MWCNTs forms a network-like structure on the surface layer of composite filter. The BET surface area of TiO2/MWCNTs/Al2O3/NF composite filter is more than 375 times higher than that of the pristine nickel foam. The degradation performance of oil particles was evaluated. Under optimized reaction conditions, 90% of the oleic acid on the filter was decomposed after 25 h. In addition, the effect of reaction temperature and light intensity were studied. Increased reaction temperature and light intensity were found to significantly improve the degradation rate and restrain the formation of refractory product. Furthermore, a parallel reaction kinetic model was adopted for degradation of oleic acid on the composite filter. The composite filter can reduces the accumulation of oil particles on the filter and the energy consumption of the filter. This approach proposes a new way to treat the cooking fume emission at residential building roofs.


Subject(s)
Air Pollutants/isolation & purification , Cooking , Nanotubes, Carbon , Titanium , Catalysis , Filtration , Kinetics , Photochemical Processes
10.
Zhongguo Yi Liao Qi Xie Za Zhi ; 42(3): 225-227, 2018 May 30.
Article in Zh | MEDLINE | ID: mdl-29885135

ABSTRACT

Distortion will occur when testing medical microwave products, due to its high power, high frequency, in the existing radiated emission testing process which will cause saturation non-linear of the pre-amplifier. It will lead to test distorted, the test results are not accurate. This article will improve the original test method by adding some suppressing-frequency devices to suppress the main frequency energy and avoid pre-amplifier saturation, eliminate distortion. All of this will ensure the test results are effective and relia ble.


Subject(s)
Equipment and Supplies , Microwaves , Amplifiers, Electronic , Equipment Design , Materials Testing
11.
Plant Cell Physiol ; 57(5): 1000-7, 2016 May.
Article in English | MEDLINE | ID: mdl-26903528

ABSTRACT

Mogrosides, the principally bioactive compounds extracted from the fruits of Siraitia grosvenorii, are a group of glycosylated cucurbitane-type tetracyclic triterpenoid saponins that exhibit a wide range of notable biological activities and are commercially available worldwide as natural sweeteners. The biosynthesis of mogrosides involves initial cyclization of 2,3-oxidosqualene to the triterpenoid skeleton of cucurbitadienol, followed by a series of oxidation reactions catalyzed by Cyt P450s (P450s) and then glycosylation reactions catalyzed by UDP glycosyltransferases (UGTs). We previously reported the identification of a cucurbitadienol synthase (SgCbQ) and a mogrol C-3 hydroxyl glycosyltransferase (UGT74AC1). However, molecular characterization of further transformation of cucurbitadienol to mogrol by P450s remains unavailable. In this study, we report the successful identification of a multifunctional P450 (CYP87D18) as being involved in C-11 oxidation of cucurbitadienol. In vitro enzymatic activity assays showed that CYP87D18 catalyzed the oxidation of cucurbitadienol at C-11 to produce 11-oxo cucurbitadienol and 11-hydroxy cucurbitadienol. Furthermore, 11-oxo-24,25-epoxy cucurbitadienol as well as 11-oxo cucurbitadienol and 11-hydroxy cucurbitadienol were produced when CYP87D18 was co-expressed with SgCbQ in genetic yeast, and their structures were confirmed by liquid chromatography-solid-phase extraction-nuclear magnetic resonance-mass spectrometry coupling (LC-SPE-NMR-MS). Taken together, these results suggest a role for CYP87D18 as a multifunctional cucurbitadienol oxidase in the mogrosides pathway.


Subject(s)
Cucurbitaceae/enzymology , Glycosides/metabolism , Plant Proteins/metabolism , Triterpenes/metabolism , Biosynthetic Pathways , Catalysis , Cucurbitaceae/genetics , Cytochrome P-450 Enzyme System/genetics , Cytochrome P-450 Enzyme System/metabolism , Fruit/enzymology , Fruit/genetics , Gene Expression , Glycosyltransferases/genetics , Glycosyltransferases/metabolism , Oxidation-Reduction , Plant Proteins/genetics , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Saponins/metabolism , Squalene/analogs & derivatives , Squalene/metabolism
12.
Electromagn Biol Med ; 35(2): 120-5, 2016.
Article in English | MEDLINE | ID: mdl-25568953

ABSTRACT

Wearable devices have been popularly used with people from different age groups. As a consequence, the concerns of their electromagnetic field (EMF) exposure to the human body and their electromagnetic interference (EMI) to the implanted medical devices have attracted many studies. The aim of this study was to evaluate the human exposure to the EMF of an active radiofrequency identification (RFID) armlet as well as its EMI to the cardiac pacemaker (CP). Different human models from various age groups were applied to assess the result variability. The scalar potential finite element method was utilized in the simulation. Local EMF exposure and the exposure to the central nerve system tissues were evaluated using different metrics. EMI to the CP was assessed in terms of the conducted voltage to the CP. The results from all the models revealed that the studied RFID armlet would not produce the EMF exposure exceeding the safety limits. The calculated interference voltage was highly dependent on the distance between the RFID armlet and the CP (i.e. the physical dimension of the individual model). The results proposed to evaluate the appropriateness of the current EMI measurement protocol for this kind of devices used by the infants.


Subject(s)
Electromagnetic Fields/adverse effects , Models, Anatomic , Pacemaker, Artificial , Radiation Exposure , Radiometry , Adult , Child , Finite Element Analysis , Humans , Infant , Male
13.
Appl Environ Microbiol ; 81(13): 4284-94, 2015 Jul.
Article in English | MEDLINE | ID: mdl-25888171

ABSTRACT

The property of loose stereochemical control at aldol products from aldolases helped to synthesize multiple polyhydroxylated compounds with nonnatural stereoconfiguration. In this study, we discovered for the first time that some fructose 1,6-diphosphate aldolases (FruA) and tagatose 1,6-diphosphate (TagA) aldolases lost their strict stereoselectivity when using l-glyceraldehyde and synthesized not only l-sorbose but also a high proportion of l-psicose. Among the aldolases tested, TagA from Bacillus licheniformis (BGatY) showed the highest enzyme activity with l-glyceraldehyde. Subsequently, a "one-pot" reaction based on BGatY and fructose-1-phosphatase (YqaB) generated 378 mg/liter l-psicose and 199 mg/liter l-sorbose from dihydroxyacetone-phosphate (DHAP) and l-glyceraldehyde. Because of the high cost and instability of DHAP, a microbial fermentation strategy was used further to produce l-sorbose/l-psicose from glucose and l-glyceraldehyde, in which DHAP was obtained from glucose through the glycolytic pathway, and some recombination pathways based on FruA or TagA and YqaB were constructed in Escherichia coli and Corynebacterium glutamicum strains. After evaluation of different host cells and combinations of FruA or TagA with YqaB and optimization of gene expression, recombinant C. glutamicum strain WT(pXFTY) was selected and produced 2.53 g/liter total ketoses, with a yield of 0.50 g/g l-glyceraldehyde. Moreover, deletion of gene cgl0331, encoding the Zn-dependent alcohol dehydrogenase in C. glutamicum, was confirmed for the first time to significantly decrease conversion of l-glyceraldehyde to glycerol and to increase yield of target products. Finally, fed-batch culture of strain SY14(pXFTY) produced 3.5 g/liter l-sorbose and 2.3 g/liter l-psicose, with a yield of 0.61 g/g l-glyceraldehyde. This microbial fermentation strategy also could be applied to efficiently synthesize other l-sugars.


Subject(s)
Aldehyde-Lyases/genetics , Aldehyde-Lyases/metabolism , Corynebacterium glutamicum/genetics , Corynebacterium glutamicum/metabolism , Fructose/biosynthesis , Metabolic Engineering/methods , Sorbose/biosynthesis , Bacillus/enzymology , Bacillus/genetics , Bacillus/metabolism , Corynebacterium glutamicum/enzymology , Escherichia coli/enzymology , Escherichia coli/genetics , Escherichia coli/metabolism , Fermentation , Gene Deletion , Glyceraldehyde/metabolism , Recombinant Proteins/genetics , Recombinant Proteins/metabolism
14.
Biotechnol Bioeng ; 112(1): 168-80, 2015 Jan.
Article in English | MEDLINE | ID: mdl-25060350

ABSTRACT

Rare sugars have various known biological functions and potential for applications in pharmaceutical, cosmetics, and food industries. Here we designed and constructed a recombination pathway in Corynebacterium glutamicum, in which dihydroxyacetone phosphate (DHAP), an intermediate of the glycolytic pathway, and a variety of aldehydes were condensed to synthesize rare ketoses sequentially by rhamnulose-1-phosphate aldolase (RhaD) and fructose-1-phosphatase (YqaB) obtained from Escherichia coli. A wild-type strain harboring this artificial pathway had the ability to produce D-sorbose and D-psicose using D-glyceraldehyde and glucose as the substrates. The tpi gene, encoding triose phosphate isomerase was further deleted, and the concentration of DHAP increased to nearly 20-fold relative to that of the wild-type. After additional optimization of expression levels from rhaD and yqaB genes and of the fermentation conditions, the engineered strain SY6(pVRTY) exhibited preferable performance for rare ketoses production. Its yield increased to 0.59 mol/mol D-glyceraldehyde from 0.33 mol/mol D-glyceraldehyde and productivity to 2.35 g/L h from 0.58 g/L h. Moreover, this strain accumulated 19.5 g/L of D-sorbose and 13.4 g/L of D-psicose using a fed-batch culture mode under the optimal conditions. In addition, it was verified that the strain SY6(pVRTY) meanwhile had the ability to synthesize C4, C5, C6, and C7 rare ketoses when a range of representative achiral and homochiral aldehydes were applied as the substrates. Therefore, the platform strain exhibited the potential for microbial production of rare ketoses and deoxysugars.


Subject(s)
Corynebacterium glutamicum/genetics , Corynebacterium glutamicum/metabolism , Ketoses/biosynthesis , Metabolic Engineering/methods , Aldehyde-Lyases/genetics , Aldehyde-Lyases/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Metabolic Networks and Pathways/genetics , Mutation , Recombination, Genetic
15.
Appl Microbiol Biotechnol ; 99(19): 7963-72, 2015 Oct.
Article in English | MEDLINE | ID: mdl-26104867

ABSTRACT

2-Deoxy-D-ribose 5-phosphate aldolase (DERA) accepts a wide variety of aldehydes and is used in de novo synthesis of 2-deoxysugars, which have important applications in drug manufacturing. However, DERA has low preference for non-phosphorylated substrates. In this study, DERA from Klebsiella pneumoniae (KDERA) was mutated to increase its enzyme activity and substrate tolerance towards non-phosphorylated polyhydroxy aldehyde. Mutant KDERA(K12) (S238D/F200I/ΔY259) showed a 3.15-fold improvement in enzyme activity and a 1.54-fold increase in substrate tolerance towards D-glyceraldehyde compared with the wild type. Furthermore, a whole-cell transformation strategy using resting cells of the BL21(pKDERA12) strain, containing the expressed plasmid pKDERA12, resulted in increase in 2-deoxy-D-ribose yield from 0.41 mol/mol D-glyceraldehyde to 0.81 mol/mol D-glyceraldehyde and higher substrate tolerance from 0.5 to 3 M compared to in vitro assays. With further optimization of the transformation process, the BL21(pKDERA12) strain produced 2.14 M (287.06 g/L) 2-deoxy-D-robose (DR), with a yield of 0.71 mol/mol D-glyceraldehyde and average productivity of 0.13 mol/L·h (17.94 g/L·h). These results demonstrate the potential for large-scale production of 2-deoxy-D-ribose using the BL21(pKDERA12) strain. Furthermore, the BL21(pKDERA12) strain also exhibited the ability to efficiently produce 2-deoxy-D-altrose from D-erythrose, as well as 2-deoxy-L-xylose and 2-deoxy-L-ribose from L-glyceraldehyde.


Subject(s)
Bacterial Proteins/metabolism , Escherichia coli/metabolism , Fructose-Bisphosphate Aldolase/metabolism , Klebsiella pneumoniae/enzymology , Ribosemonophosphates/biosynthesis , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Catalysis , Cloning, Molecular , Escherichia coli/genetics , Fructose-Bisphosphate Aldolase/chemistry , Fructose-Bisphosphate Aldolase/genetics , Gene Expression , Kinetics , Substrate Specificity
16.
J Ind Microbiol Biotechnol ; 42(5): 661-9, 2015 May.
Article in English | MEDLINE | ID: mdl-25724336

ABSTRACT

An engineered strain for the conversion of D-fructose to allitol was developed by constructing a multi-enzyme coupling pathway and cofactor recycling system in Escherichia coli. D-Psicose-3-epimerase from Ruminococcus sp. and ribitol dehydrogenase from Klebsiella oxytoca were coexpressed to form the multi-enzyme coupling pathway for allitol production. The cofactor recycling system was constructed using the formate dehydrogenase gene from Candida methylica for continuous NADH supply. The recombinant strain produced 10.62 g/l allitol from 100 mM D-fructose. To increase the intracellular concentration of the substrate, the glucose/fructose facilitator gene from Zymomonas mobilis was incorporated into the engineered strain. The results showed that the allitol yield was enhanced significantly to 16.53 g/l with a conversion rate of 92 %. Through optimizing conversion conditions, allitol was produced effectively on a large scale by the whole-cell biotransformation system; the yield reached 48.62 g/l when 500 mM D-fructose was used as the substrate.


Subject(s)
Biosynthetic Pathways/genetics , Carbohydrate Epimerases/genetics , Escherichia coli/genetics , Formate Dehydrogenases/genetics , Metabolic Engineering , Sugar Alcohol Dehydrogenases/genetics , Sugar Alcohols/metabolism , Bioreactors , Biotransformation , Candida/enzymology , Candida/genetics , Carbohydrate Epimerases/metabolism , Escherichia coli/metabolism , Formate Dehydrogenases/metabolism , Fructose/metabolism , Klebsiella oxytoca/enzymology , Klebsiella oxytoca/genetics , NAD/metabolism , Racemases and Epimerases/genetics , Racemases and Epimerases/metabolism , Ruminococcus/enzymology , Ruminococcus/genetics , Sugar Alcohol Dehydrogenases/metabolism , Zymomonas/genetics
17.
Biotechnol Lett ; 36(3): 553-60, 2014 Mar.
Article in English | MEDLINE | ID: mdl-24129953

ABSTRACT

A dual route for anaerobic succinate production was engineered into Corynebacterium glutamicum. The glyoxylate pathway was reconstructed by overexpressing isocitrate lyase, malate synthase and citrate synthase. The engineered strain produced succinate with a yield of 1.34 mol (mol glucose)(-1). Further overexpression of succinate exporter, SucE, increased succinate yield to 1.43 mol (mol glucose)(-1). Metabolic flux analysis revealed that the glyoxylate pathway was further activated by engineering succinate export system. Using an anaerobic fed-batch fermentation process, the final strain produced 926 mM succinate (= 109 g l(-1)) with an overall volumetric productivity of 9.4 mM h(-1) and an average yield of 1.32 mol (mol glucose)(-1).


Subject(s)
Corynebacterium glutamicum/genetics , Corynebacterium glutamicum/metabolism , Metabolic Engineering/methods , Metabolic Networks and Pathways/genetics , Succinic Acid/metabolism , Anaerobiosis , Batch Cell Culture Techniques , Citrate (si)-Synthase/genetics , Citrate (si)-Synthase/metabolism , Fermentation , Gene Expression , Glyoxylates/metabolism , Isocitrate Lyase/genetics , Isocitrate Lyase/metabolism , Malate Synthase/genetics , Malate Synthase/metabolism , Metabolic Flux Analysis
18.
J Basic Microbiol ; 54(10): 1073-9, 2014 Oct.
Article in English | MEDLINE | ID: mdl-24771547

ABSTRACT

A novel bacterium capable of producing allitol from D-psicose was isolated from soil and identified as Klebsiella oxytoca G4A4. An efficient method for the transformation of D-psicose to allitol was achieved through the resting cell reaction. Ribitol as an inducer is suitable for cell cultivation, and cells are most active in Tris-HCl buffer (pH 8.0) at 37 °C with a density of 40 (OD600 nm ). After the reaction, the final conversion rates of the washed cells were approximately 87, 83, and 55% at D-psicose concentrations of 0.25, 0.5, and 1%, respectively. The product from D-psicose was purified and determined to be allitol by high-performance liquid chromatography and nuclear magnetic resonance spectroscopy.


Subject(s)
Fructose/metabolism , Klebsiella oxytoca/metabolism , Sugar Alcohols/metabolism , Chromatography, High Pressure Liquid , Klebsiella oxytoca/isolation & purification , Magnetic Resonance Spectroscopy , Soil Microbiology
19.
Zhongguo Yi Liao Qi Xie Za Zhi ; 38(6): 430-2, 2014 Nov.
Article in Zh | MEDLINE | ID: mdl-25980132

ABSTRACT

With the enforcement of YY 0505-2012, the testing items and evaluation points of radio frequency ablation catheter in electromagnetic compatibility field should be studied and discussed. Based on the requirements of relevant standards, this paper discusses on the testing items that should be applied and the evaluation points that should be focused on by analyzing the intended use and the structure of radio frequency ablation catheter, when it intends to apply registration individually with the basic knowledge of electromagnetic compatibility field.


Subject(s)
Catheter Ablation/instrumentation , Electromagnetic Fields , Electromagnetic Phenomena , Evaluation Studies as Topic
20.
Article in English | MEDLINE | ID: mdl-39017714

ABSTRACT

Benign prostatic hyperplasia (BPH) is a prevalent urological condition that predominantly affects the geriatric male population, resulting in lower urinary tract symptoms. Saw palmetto is a traditional Chinese medicine for treating BPH. This study aimed to investigate the potential therapeutic mechanisms of saw palmetto in BPH treatment. The active ingredients and potential targets of saw palmetto were obtained through the TCMSP database. BPH-related targets were retrieved from the GeneCards database. PPI, GO, and KEEG analyses were performed to predict the potential therapeutic mechanism. The active ingredient-common target and common target-pathway networks were constructed by Cytoscape software. Molecular docking and cellular experiments were carried out to further validate the potential mechanism. We obtained 13 active components in saw palmetto and 56 common targets in BPH treatment. KEEG analysis showed that the estrogen signaling pathway was the most enriched and exhibited a close association with BPH. PPI analysis, along with ingredient-target and target-pathway network analyses, indicated that stigmasterol was the core ingredient and PGR, NCOA1, and NCOA2 were identified as the hub genes mediating the effects of saw palmetto against BPH. In addition, molecular docking showed that stigmasterol had strong binding to PGR, NCOA1, and NCOA2. Cellular experiments revealed that stigmasterol significantly increased the percentage of BPH-1 cells in the G0/G1 phase and inhibited cell viability and division. Furthermore, it notably reduced the expression of PGR, NCOA1, and NCOA2. Saw palmetto might inhibit cell viability and division by suppressing the expression of PGR, NCOA1, and NCOA2, thereby playing a therapeutic role in treating BPH.

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