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1.
Mol Biol Evol ; 40(8)2023 08 03.
Artigo em Inglês | MEDLINE | ID: mdl-37541261

RESUMO

Centromeres (CEN) are the chromosomal regions that play a crucial role in maintaining genomic stability. The underlying highly repetitive DNA sequences can evolve quickly in most eukaryotes, and promote karyotype evolution. Despite their variability, it is not fully understood how these widely variable sequences ensure the homeostasis of centromere function. In this study, we investigated the genetics and epigenetics of CEN in a population of wheat lines from global breeding programs. We captured a high degree of sequences, positioning, and epigenetic variations in the large and complex wheat CEN. We found that most CENH3-associated repeats are Cereba element of retrotransposons and exhibit phylogenetic homogenization across different wheat lines, but the less-associated repeat sequences diverge on their own way in each wheat line, implying specific mechanisms for selecting certain repeat types as functional core CEN. Furthermore, we observed that CENH3 nucleosome structures display looser wrapping of DNA termini on complex centromeric repeats, including the repositioned CEN. We also found that strict CENH3 nucleosome positioning and intrinsic DNA features play a role in determining centromere identity among different lines. Specific non-B form DNAs were substantially associated with CENH3 nucleosomes for the repositioned centromeres. These findings suggest that multiple mechanisms were involved in the adaptation of CENH3 nucleosomes that can stabilize CEN. Ultimately, we proposed a remarkable epigenetic plasticity of centromere chromatin within the diverse genomic context, and the high robustness is crucial for maintaining centromere function and genome stability in wheat 10+ lines as a result of past breeding selections.


Assuntos
Histonas , Nucleossomos , Histonas/genética , Triticum/genética , Filogenia , Melhoramento Vegetal , Centrômero/genética
2.
Metab Eng ; 82: 100-109, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38325640

RESUMO

Odd-numbered fatty acids (FAs) have been widely used in nutrition, agriculture, and chemical industries. Recently, some studies showed that they could be produced from bacteria or yeast, but the products are almost exclusively odd-numbered long-chain FAs. Here we report the design and construction of two biosynthetic pathways in Saccharomyces cerevisiae for de novo production of odd-numbered medium-chain fatty acids (OMFAs) via ricinoleic acid and 10-hydroxystearic acid, respectively. The production of OMFAs was enabled by introducing a hydroxy fatty acid cleavage pathway, including an alcohol dehydrogenase from Micrococcus luteus, a Baeyer-Villiger monooxygenase from Pseudomonas putida, and a lipase from Pseudomonas fluorescens. These OMFA biosynthetic pathways were optimized by eliminating the rate-limiting step, generating heptanoic acid, 11-hydroxyundec-9-enoic acid, nonanoic acid, and 9-hydroxynonanoic acid at 7.83 mg/L, 9.68 mg/L, 9.43 mg/L and 13.48 mg/L, respectively. This work demonstrates the biological production of OMFAs in a sustainable manner in S. cerevisiae.


Assuntos
Engenharia Metabólica , Saccharomyces cerevisiae , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Ácidos Graxos , Oxigenases de Função Mista/metabolismo , Álcool Desidrogenase/metabolismo
3.
Sensors (Basel) ; 24(13)2024 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-39001006

RESUMO

Infrared small target detection technology plays a crucial role in various fields such as military reconnaissance, power patrol, medical diagnosis, and security. The advancement of deep learning has led to the success of convolutional neural networks in target segmentation. However, due to challenges like small target scales, weak signals, and strong background interference in infrared images, convolutional neural networks often face issues like leakage and misdetection in small target segmentation tasks. To address this, an enhanced U-Net method called MST-UNet is proposed, the method combines multi-scale feature decomposition and fusion and attention mechanisms. The method involves using Haar wavelet transform instead of maximum pooling for downsampling in the encoder to minimize feature loss and enhance feature utilization. Additionally, a multi-scale residual unit is introduced to extract contextual information at different scales, improving sensory field and feature expression. The inclusion of a triple attention mechanism in the encoder structure further enhances multidimensional information utilization and feature recovery by the decoder. Experimental analysis on the NUDT-SIRST dataset demonstrates that the proposed method significantly improves target contour accuracy and segmentation precision, achieving IoU and nIoU values of 80.09% and 80.19%, respectively.

4.
Int J Mol Sci ; 25(12)2024 Jun 08.
Artigo em Inglês | MEDLINE | ID: mdl-38928068

RESUMO

As a low-calorie sugar, D-allulose is produced from D-fructose catalyzed by D-allulose 3-epimerase (DAE). Here, to improve the catalytic activity, stability, and processability of DAE, we reported a novel method by forming organic-inorganic hybrid nanoflowers (NF-DAEs) and co-immobilizing them on resins to form composites (Re-NF-DAEs). NF-DAEs were prepared by combining DAE with metal ions (Co2+, Cu2+, Zn2+, Ca2+, Ni2+, Fe2+, and Fe3+) in PBS buffer, and were analyzed by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy, and X-ray diffraction. All of the NF-DAEs showed higher catalytic activities than free DAE, and the NF-DAE with Ni2+ (NF-DAE-Ni) reached the highest relative activity of 218%. The NF-DAEs improved the thermal stability of DAE, and the longest half-life reached 228 min for NF-DAE-Co compared with 105 min for the free DAE at 55 °C. To further improve the recycling performance of the NF-DAEs in practical applications, we combined resins and NF-DAEs to form Re-NF-DAEs. Resins and NF-DAEs co-effected the performance of the composites, and ReA (LXTE-606 neutral hydrophobic epoxy-based polypropylene macroreticular resins)-based composites (ReA-NF-DAEs) exhibited outstanding relative activities, thermal stabilities, storage stabilities, and processabilities. The ReA-NF-DAEs were able to be reused to catalyze the conversion from D-fructose to D-allulose, and kept more than 60% of their activities after eight cycles.


Assuntos
Estabilidade Enzimática , Enzimas Imobilizadas , Enzimas Imobilizadas/química , Carboidratos Epimerases/química , Carboidratos Epimerases/metabolismo , Nanoestruturas/química , Frutose/química , Espectroscopia de Infravermelho com Transformada de Fourier , Difração de Raios X
5.
Biotechnol Bioeng ; 120(3): 852-858, 2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36464776

RESUMO

Odd-chain fatty acids (OCFAs) and their derivatives have attracted increasing attention due to their wide applications in the chemical, fuel, and pharmaceutical industry. However, most natural fatty acids are even-chained, and OCFAs are rare. In this work, a novel pathway was designed and established for de novo synthesis of OCFAs via 3-hydroxypropionic acid (3-HP) as the intermediate in Saccharomyces cerevisiae. First, the OCFAs biosynthesis pathway from 3-HP was confirmed, followed by an optimization of the precursor 3-HP. After combining these strategies, a de novo production of OCFAs at 74.8 mg/L was achieved, and the percentage of OCFAs in total lipids reached 20.3%, reaching the highest ratio of de novo-produced OCFAs. Of the OCFAs produced by the engineered strain, heptadecenoic acid (C17:1) and heptadecanoic acid (C17:0) accounted for 12.1% and 7.6% in total lipid content, respectively. This work provides a new and promising pathway for the de novo bio-production of OCFAs.


Assuntos
Ácidos Graxos , Saccharomyces cerevisiae , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Ácidos Graxos/metabolismo , Ácidos Graxos Monoinsaturados/metabolismo , Ácido Láctico/metabolismo , Engenharia Metabólica
6.
Sensors (Basel) ; 23(15)2023 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-37571539

RESUMO

Convolutional neural networks have achieved good results in target detection in many application scenarios, but convolutional neural networks still face great challenges when facing scenarios with small target sizes and complex background environments. To solve the problem of low accuracy of infrared weak target detection in complex scenes, and considering the real-time requirements of the detection task, we choose the YOLOv5s target detection algorithm for improvement. We add the Bottleneck Transformer structure and CoordConv to the network to optimize the model parameters and improve the performance of the detection network. Meanwhile, a two-dimensional Gaussian distribution is used to describe the importance of pixel points in the target frame, and the normalized Guassian Wasserstein distance (NWD) is used to measure the similarity between the prediction frame and the true frame to characterize the loss function of weak targets, which will help highlight the targets with flat positional deviation transformation and improve the detection accuracy. Finally, through experimental verification, compared with other mainstream detection algorithms, the improved algorithm in this paper significantly improves the target detection accuracy, with the mAP reaching 96.7 percent, which is 2.2 percentage points higher compared with Yolov5s.

7.
Ann Hepatol ; 27(3): 100680, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35108614

RESUMO

INTRODUCTION AND OBJECTIVES: Hepatocellular carcinoma (HCC) is one of the most malignant digestive tumors, and its insidious onset and rapid progression are the main reasons for the difficulty in effective treatment. Lysophosphatidylcholine acyltransferase 1 (LPCAT1) is a key enzyme that regulates phospholipid metabolism of the cell membrane. However, the mechanism by which LPCAT1 regulates HCC metastasis remains unknown. This study aimed to explore its biological function and potential mechanisms concerning migration and invasion in HCC. MATERIALS AND METHODS: LPCAT1 expression in HCC tissues and its association with clinical outcomes were investigated by western blotting and bioinformatic methods, respectively. The role of LPCAT1 in migration and invasion was assessed via Transwell assays. The expression pattern of epithelial-mesenchymal transition (EMT) markers was quantified by western blotting. The biological behaviors of LPCAT1 in vivo were evaluated using xenograft tumor models and caudal vein metastatic models. Signaling pathways related to LPCAT1 were predicted using gene set enrichment analysis (GSEA) and further confirmed by western blotting. RESULTS: LPCAT1 expression was significantly upregulated in HCC tissues and indicated a poor prognosis of HCC patients. Several EMT-related markers were found to be regulated by LPCAT1. HCC cells overexpressing LPCAT1 exhibited remarkably high migration and invasion capacities, upregulated expression of mesenchymal markers and reduced E-cadherin expression. In vivo, LPCAT1 promoted HCC pulmonary metastasis. Furthermore, the Wnt/ß-catenin signaling pathway was confirmed to be activated by LPCAT1. CONCLUSIONS: LPCAT1 could serve as a promising biomarker of HCC and as a novel therapeutic target for the treatment of metastatic HCC.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , 1-Acilglicerofosfocolina O-Aciltransferase/genética , 1-Acilglicerofosfocolina O-Aciltransferase/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Transição Epitelial-Mesenquimal , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/patologia , Via de Sinalização Wnt/genética
8.
Entropy (Basel) ; 24(9)2022 Sep 11.
Artigo em Inglês | MEDLINE | ID: mdl-36141164

RESUMO

In this paper, we focus on the nonsmooth composite optimization problems over networks, which consist of a smooth term and a nonsmooth term. Both equality constraints and box constraints for the decision variables are also considered. Based on the multi-agent networks, the objective problems are split into a series of agents on which the problems can be solved in a decentralized manner. By establishing the Lagrange function of the problems, the first-order optimal condition is obtained in the primal-dual domain. Then, we propose a decentralized algorithm with the proximal operators. The proposed algorithm has uncoordinated stepsizes with respect to agents or edges, where no global parameters are involved. By constructing the compact form of the algorithm with operators, we complete the convergence analysis with the fixed-point theory. With the constrained quadratic programming problem, simulations verify the effectiveness of the proposed algorithm.

9.
Pharmacol Res ; 165: 105464, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33515707

RESUMO

BACKGROUND: An individual's level of lower limb motor function is associated with his or her disability level after stroke, and motor improvement may lead to a better prognosis and quality of life. Data from animal models show that Qizhitongluo (QZTL) capsule facilitates recovery after focal brain injury. We aimed to validate the efficacy and safety of the QZTL capsule for promoting lower limb motor recovery in poststroke patients. METHODS: In this randomized, multicenter, double-blind, placebo- and active-controlled trial from 13 sites in China, participants with ischemic stroke and Fugl-Meyer motor scale (FMMS) scores of <95 were eligible for inclusion. Patients were randomly assigned in a 2:1:1 ratio to the QZTL group, Naoxintong (NXT) group or placebo group for 12 weeks at 15-28 days after the onset of stroke. The primary outcome was the change in the Lower Limb FMMS (FMMS-LL) score from baseline over the 12-week intervention period. RESULTS: 622 participants were randomly assigned to the QZTL group (309), NXT group (159), or placebo group (154). The FMMS-LL score increased by 4.81 points (95 % CI, 4.27-5.35) in the QZTL group, by 3.77 points (95 % CI, 3.03-4.51) in the NXT group and by 3.00 points (95 % CI, 3.03-4.51) in the placebo group at week 12. The QZTL group showed significantly larger improvements compared with the placebo group at each interview from weeks 4-12 (difference, 0.89 [0.30,1.49] at week 4, P = 0.0032; difference, 1.83[1.01,2.66] at 90 days poststroke, P < 0.0001; difference, 1.81[0.88,2.74] at week 12, P = 0.0001). CONCLUSION: The QZTL capsule is an effective treatment for lower limb motor impairment. The finding indicates that the QZTL capsule may be used as a potential new strategy for stroke rehabilitation.


Assuntos
Medicamentos de Ervas Chinesas/uso terapêutico , Extremidade Inferior/fisiologia , Reabilitação do Acidente Vascular Cerebral/métodos , Acidente Vascular Cerebral/diagnóstico , Acidente Vascular Cerebral/terapia , Idoso , Cápsulas , Método Duplo-Cego , Medicamentos de Ervas Chinesas/farmacologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Recuperação de Função Fisiológica/efeitos dos fármacos , Recuperação de Função Fisiológica/fisiologia , Acidente Vascular Cerebral/fisiopatologia , Resultado do Tratamento
10.
BMC Complement Altern Med ; 19(1): 370, 2019 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-31842860

RESUMO

BACKGROUND: Tianshu capsule (TSC), a formula of traditional Chinese medicine, has been widely used in clinical practice for prophylactic treatment of headaches in China. However, former clinical trials of TSC were small, and lack of a standard set of diagnostic criteria to enroll patients. The study was conducted to re-evaluate the efficacy and safety of TSC post-marketing in an extending number of migraineurs who have diagnosed migraine with the International Classification of Headache Disorders, 3rd edition (beta version, ICHD-3ß). METHODS: The study was a double-blind, randomized, placebo-controlled clinical trial that conducted at 20 clinical centers in China. At enrollment, patients between 18 and 65 years of age diagnosed with migraine were assigned to receive either TSC (4.08 g, three times daily) or a matched placebo according to a randomization protocol. The primary endpoint was a relative reduction of 50% or more in the frequency of headache attacks. The secondary outcomes included a reduction in the incidence of headache, the visual analogue scale of headache attacks, days of acute analgesic usage, and percentage of patients with a decrease of 50% or more in headache severity. Accompanying symptoms were also assessed. RESULTS: One thousand migraine patients were initially enrolled in the study, and 919 of them completed the trial. Following the 12-week treatment, significant improvement was observed in the TSC group concerning both primary and secondary outcomes. After therapy discontinuation, the gap between the TSC group and the placebo group in efficacy outcomes continued to increase. There were no severe adverse effects. CONCLUSIONS: TSC is an effective, well-tolerated medicine for prophylactic treatment of migraine, and still have prophylactic effect after medicine discontinuation. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02035111; Data of registration: 2014-01-10.


Assuntos
Analgésicos/uso terapêutico , Medicamentos de Ervas Chinesas/uso terapêutico , Transtornos de Enxaqueca/tratamento farmacológico , Adulto , Analgésicos/efeitos adversos , Método Duplo-Cego , Medicamentos de Ervas Chinesas/efeitos adversos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Resultado do Tratamento
11.
Microb Cell Fact ; 16(1): 125, 2017 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-28724386

RESUMO

Plant natural products (PNPs) are widely used as pharmaceuticals, nutraceuticals, seasonings, pigments, etc., with a huge commercial value on the global market. However, most of these PNPs are still being extracted from plants. A resource-conserving and environment-friendly synthesis route for PNPs that utilizes microbial cell factories has attracted increasing attention since the 1940s. However, at the present only a handful of PNPs are being produced by microbial cell factories at an industrial scale, and there are still many challenges in their large-scale application. One of the challenges is that most biosynthetic pathways of PNPs are still unknown, which largely limits the number of candidate PNPs for heterologous microbial production. Another challenge is that the metabolic fluxes toward the target products in microbial hosts are often hindered by poor precursor supply, low catalytic activity of enzymes and obstructed product transport. Consequently, despite intensive studies on the metabolic engineering of microbial hosts, the fermentation costs of most heterologously produced PNPs are still too high for industrial-scale production. In this paper, we review several aspects of PNP production in microbial cell factories, including important design principles and recent progress in pathway mining and metabolic engineering. In addition, implemented cases of industrial-scale production of PNPs in microbial cell factories are also highlighted.


Assuntos
Bactérias/metabolismo , Produtos Biológicos , Engenharia Celular , Fungos/metabolismo , Microbiologia Industrial , Preparações de Plantas/isolamento & purificação , Bactérias/genética , Vias Biossintéticas , Suplementos Nutricionais , Fermentação , Fungos/genética , Engenharia Metabólica , Preparações de Plantas/química , Preparações de Plantas/metabolismo , Preparações de Plantas/uso terapêutico , Biologia Sintética
12.
Microb Cell Fact ; 16(1): 165, 2017 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-28950867

RESUMO

BACKGROUND: Kaempferol is a flavonol with broad bioactivity of anti-oxidant, anti-cancer, anti-diabetic, anti-microbial, cardio-protective and anti-asthma. Microbial synthesis of kaempferol is a promising strategy because of the low content in primary plant source. METHODS: In this study, the biosynthesis pathway of kaempferol was constructed in the budding yeast Saccharomyces cerevisiae to produce kaempferol de novo, and several biological measures were taken for high production. RESULTS: Firstly, a high efficient flavonol synthases (FLS) from Populus deltoides was introduced into the biosynthetic pathway of kaempferol. Secondly, a S. cerevisiae recombinant was constructed for de novo synthesis of kaempferol, which generated about 6.97 mg/L kaempferol from glucose. To further promote kaempferol production, the acetyl-CoA biosynthetic pathway was overexpressed and p-coumarate was supplied as substrate, which improved kaempferol titer by about 23 and 120%, respectively. Finally, a fed-batch process was developed for better kaempferol fermentation performance, and the production reached 66.29 mg/L in 40 h. CONCLUSIONS: The titer of kaempferol in our engineered yeast is 2.5 times of the highest reported titer. Our study provides a possible strategy to produce kaempferol using microbial cell factory.


Assuntos
Quempferóis/biossíntese , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Vias Biossintéticas , Fermentação , Glucose/metabolismo , Engenharia Metabólica
13.
J Colloid Interface Sci ; 671: 15-33, 2024 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-38788421

RESUMO

The exploration of bifunctional electrocatalysts with high activity, stability, and economy is of great significance in promoting the development of water splitting. Herein, a dual active sites heterostructure NiCoS/NC was designed to be derived in situ on 3D N-doped porous carbon (NC) using gelatin as a nitrogen and carbon source. The characterization of experiments suggests that nanoflower-like Ni2CoS4 (abbreviated as NiCoS) was randomly distributed on the NC substrate, and the sheet-like NC formed a highly open porous network structure resembling a honeycomb, which provided more accessible active sites for electrolyte ions. In addition, the special nanostructures of the catalyst materials help to promote the surface reconstruction to the real active substance NiOOH/CoOOH, and the double active sites synergistically reduce the overpotential of OER and improve its kinetics. DFT (Density-functional theory) calculations reveal the electronic coupling of NiCoS/NC in atomic orbitals, modulation of electrons by the heterointerface and N-doping, and synergistic effect of dual active sites improving the inherent catalytic activity. The NiCoS/NC composite electrocatalyst exhibited a 177 mV small OER overpotential and a 132 mV small HER overpotential with Faraday efficiencies as high as 96 % and 98 % at 10 mA cm-2 current density. In the two-electrode system, it also requires only an ultra-low voltage of 1.52 V to achieve a 10 mA cm-2 current density, and it shows excellent long-term water splitting stability. This provides a new idea for the development of transition metal-based bifunctional electrocatalysts.

14.
Int J Biol Macromol ; 259(Pt 2): 129369, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38218271

RESUMO

The impact of the cell wall structure of Monascus purpureus M9 on the secretion of extracellular monascus pigments (exMPs) was investigated. To modify the cell wall structure, UDP-galactopyranose mutase (GlfA) was knocked out using Agrobacterium-mediated transformation method, leading to a significant reduction in the Galf-based polysaccharide within the cell wall. Changes in mycelium morphology, sporogenesis, and the expression of relevant genes in M9 were also observed following the mutation. Regarding MPs secretion, a notable increase was observed in six types of exMPs (R1, R2, Y1, Y2, O1 and O2). Specifically, these exMPs exhibited enhancement of 1.33, 1.59, 0.8, 2.45, 2.89 and 4.03 times, respectively, compared to the wild-type strain. These findings suggest that the alteration of the cell wall structure could selectively influence the secretion of MPs in M9. The underlying mechanisms were also discussed. This research contributes new insights into the regulation of the synthesis and secretion of MPs in Monascus spp..


Assuntos
Galactose/análogos & derivados , Transferases Intramoleculares , Mananas , Monascus , Monascus/genética , Monascus/metabolismo , Pigmentos Biológicos , Metabolismo dos Carboidratos , Fermentação
15.
Appl Environ Microbiol ; 79(10): 3273-81, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23503313

RESUMO

Glycerol is a major by-product in bioethanol fermentation by the yeast Saccharomyces cerevisiae, and decreasing glycerol formation for increased ethanol yield has been a major research effort in the bioethanol field. A new strategy has been used in the present study for reduced glycerol formation and improved ethanol fermentation performance by finely modulating the expression of GPD1 in the KAM15 strain (fps1Δ pPGK1-GLT1 gpd2Δ). The GPD1 promoter was serially truncated from the 3' end by 20 bp to result in a different expression strength of GPD1. The two engineered promoters carrying 60- and 80-bp truncations exhibited reduced promoter strength but unaffected osmostress response. These two promoters were integrated into the KAM15 strain, generating strains LE34U and LE35U, respectively. The transcription levels of LE34U and LE35U were 37.77 to 45.12% and 21.34 to 24.15% of that of KAM15U, respectively, depending on osmotic stress imposed by various glucose concentrations. In very high gravity (VHG) fermentation, the levels of glycerol for LE34U and LE35U were reduced by 15.81% and 30.66%, respectively, compared to KAM15U. The yield and final concentration of ethanol for LE35U were 3.46% and 0.33% higher, respectively, than those of KAM15U. However, fermentation rate and ethanol productivity for LE35U were reduced. On the other hand, the ethanol yield and final concentration for LE34U were enhanced by 2.28% and 2.32%, respectively, compared to those of KAM15U. In addition, a 2.31% increase in ethanol productivity was observed for LE34U compared to KAM15U. These results verified the feasibility of our strategy for yeast strain development.


Assuntos
Etanol/metabolismo , Glicerol-3-Fosfato Desidrogenase (NAD+)/genética , Regiões Promotoras Genéticas , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Deleção de Sequência , Adaptação Biológica , Sequência de Bases , Fermentação , Engenharia Genética/métodos , Glicerol/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crescimento & desenvolvimento , Estresse Fisiológico , Transcrição Gênica , Transformação Genética
16.
Biotechnol Lett ; 35(11): 1859-64, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23801122

RESUMO

We have investigated whether simultaneous modification of cofactor metabolism and glycerol in a strain of Saccharomyces cerevisiae can eliminate glycerol synthesis during ethanol production. Two strains, S812 (gpd1Δ gpd2Δ PGK1p-GLT1) and LE17 (gpd1Δ gpd2Δ PGK1p-GLT1 PGKp-STL1) were generated that showed a 8 and 8.2 % increase in the ethanol yield, respectively, compared to the wild type KAM-2 strain. The ethanol titer was improved from 90.4 g/l for KAM-2 to 97.6 g/l for S812 and 97.8 g/l for LE17, respectively. These results provide a new insight into rationalization of metabolic engineering strategies for improvement of ethanol yield through elimination of glycerol production.


Assuntos
Etanol/metabolismo , Glutamato Sintase/biossíntese , Glicerol-3-Fosfato Desidrogenase (NAD+)/deficiência , Proteínas de Membrana Transportadoras/biossíntese , Engenharia Metabólica , Proteínas de Saccharomyces cerevisiae/biossíntese , Saccharomyces cerevisiae/enzimologia , Etanol/toxicidade , Fermentação , Glicerol/metabolismo , Saccharomyces cerevisiae/efeitos dos fármacos
17.
Appl Environ Microbiol ; 78(4): 1081-6, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22156411

RESUMO

The recombinant xylose-fermenting Saccharomyces cerevisiae strain harboring xylose reductase (XR) and xylitol dehydrogenase (XDH) from Scheffersomyces stipitis requires NADPH and NAD(+), creates cofactor imbalance, and causes xylitol accumulation during growth on d-xylose. To solve this problem, noxE, encoding a water-forming NADH oxidase from Lactococcus lactis driven by the PGK1 promoter, was introduced into the xylose-utilizing yeast strain KAM-3X. A cofactor microcycle was set up between the utilization of NAD(+) by XDH and the formation of NAD(+) by water-forming NADH oxidase. Overexpression of noxE significantly decreased xylitol formation and increased final ethanol production during xylose fermentation. Under xylose fermentation conditions with an initial d-xylose concentration of 50 g/liter, the xylitol yields for of KAM-3X(pPGK1-noxE) and control strain KAM-3X were 0.058 g/g xylose and 0.191 g/g, respectively, which showed a 69.63% decrease owing to noxE overexpression; the ethanol yields were 0.294 g/g for KAM-3X(pPGK1-noxE) and 0.211 g/g for the control strain KAM-3X, which indicated a 39.33% increase due to noxE overexpression. At the same time, the glycerol yield also was reduced by 53.85% on account of the decrease in the NADH pool caused by overexpression of noxE.


Assuntos
Complexos Multienzimáticos/metabolismo , NADH NADPH Oxirredutases/metabolismo , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/metabolismo , Xilitol/metabolismo , Xilose/metabolismo , Etanol/metabolismo , Fermentação , Expressão Gênica , Lactococcus lactis/enzimologia , Lactococcus lactis/genética , Engenharia Metabólica/métodos , Complexos Multienzimáticos/genética , NADH NADPH Oxirredutases/genética , Regiões Promotoras Genéticas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Água/metabolismo
18.
Biotechnol J ; 17(3): e2100579, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35086163

RESUMO

BACKGROUND: Propionate is widely used as a preservative in the food and animal feed industries. Propionate is currently produced by petrochemical processes, and fermentative production of propionate remains challenging. METHODS AND RESULTS: In this study, a synthetic propionate pathway was constructed in the budding yeast Saccharomyces cerevisiae, for propionate production under aerobic conditions. Through expression of tdcB and aldH from Escherichia coli and kivD from Lactococcus lactis, L-threonine was converted to propionate via 2-ketobutyrate and propionaldehyde. The resulting yeast aerobically produced 0.21 g L-1 propionate from glucose in a shake flask. Subsequent overexpression of pathway genes and elimination of competing pathways increased propionate production to 0.37 g L-1 . To further increase propionate production, carbon flux was pulled into the propionate pathway by weakened expression of pyruvate kinase (PYK1), together with overexpression of phosphoenolpyruvate carboxylase (ppc). The final propionate production reached 1.05 g L-1 during fed-batch fermentation in a fermenter. CONCLUSIONS AND IMPLICATIONS: In this work, a yeast cell factory was constructed using synthetic biology and metabolic engineering strategies to enable propionate production under aerobic conditions. Our study demonstrates engineered S. cerevisiae as a promising alternative for the production of propionate and its derivatives.


Assuntos
Engenharia Metabólica , Saccharomyces cerevisiae , Fermentação , Engenharia Metabólica/métodos , Propionatos/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Treonina/genética , Treonina/metabolismo
19.
Appl Microbiol Biotechnol ; 91(4): 1239-46, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21706172

RESUMO

Genome shuffling is an efficient way to improve complex phenotypes under the control of multiple genes. For the improvement of strain's performance in very high-gravity (VHG) fermentation, we developed a new method of genome shuffling. A diploid ste2/ste2 strain was subjected to EMS (ethyl methanesulfonate) mutagenesis followed by meiotic recombination-mediated genome shuffling. The resulting haploid progenies were intrapopulation sterile and therefore haploid recombinant cells with improved phenotypes were directly selected under selection condition. In VHG fermentation, strain WS1D and WS5D obtained by this approach exhibited remarkably enhanced tolerance to ethanol and osmolarity, increased metabolic rate, and 15.12% and 15.59% increased ethanol yield compared to the starting strain W303D, respectively. These results verified the feasibility of the strain improvement strategy and suggested that it is a powerful and high throughput method for development of Saccharomyces cerevisiae strains with desired phenotypes that is complex and cannot be addressed with rational approaches.


Assuntos
Etanol/metabolismo , Redes e Vias Metabólicas/genética , Mutagênese , Recombinação Genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Embaralhamento de DNA , Metanossulfonato de Etila/metabolismo , Fermentação , Mutagênicos/metabolismo
20.
Zhonghua Wai Ke Za Zhi ; 49(6): 522-5, 2011 Jun 01.
Artigo em Zh | MEDLINE | ID: mdl-21914302

RESUMO

OBJECTIVE: To evaluate the safety and efficacy of transanal drainage tube followed by laparoscopic surgery in management of malignant colorectal obstruction. METHODS: From March 2007 to October 2010, 37 patients with colorectal cancer manifesting acute complete mechanical obstruction were treated by ileus tube drainage. After irrigation and drainage ranging from 4 to 10 days, the radical operations and anastomosis were performed by laparoscopy. RESULTS: The drainage tubes were successfully implanted in 34 patients. The decompression time of patients was (5.8 ± 1.6) d, ranging from 4 to 10 d. The abdominal pain and bloating symptoms were faded away after (3.8 ± 1.3) d (1 to 7 d) drainage. And comparing to that of patients when admission, abdominal circumference significantly reduced from (92 ± 7) cm to (84 ± 6) cm (P = 0.013) before surgery. Thirty-one cases were performed radical resection and anastomosis by laparoscopy after decompression. Postoperative recovery was smooth, and there was no serious complication. CONCLUSIONS: Laparoscopic surgery followed decompression by transanal ileus tube is effective and safe for acute lower colorectal obstruction. Emergency surgery may be converted to limit surgery by this method. After appropriate bowel preparation, laparoscopic radical surgery and anastomosis is feasible.


Assuntos
Neoplasias Colorretais/cirurgia , Drenagem/métodos , Obstrução Intestinal/cirurgia , Laparoscopia , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Colorretais/patologia , Feminino , Humanos , Obstrução Intestinal/etiologia , Masculino , Pessoa de Meia-Idade
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