Combinatorial Fine-Tuning of Phospholipase D Expression by Bacillus subtilis WB600 for the Production of Phosphatidylserine

( Tingting Huang ),( Xueqin Lv ),( Jianghua Li ),( Hyun-dong Shin ),( Guocheng Du ),( Long Liu ) 한국미생물 · 생명공학회 2018 Journal of microbiology and biotechnology Vol.28 No.12

Phospholipase D has great commercial value due to its transphosphatidylation products that can be used in the food and medicine industries. In order to construct a strain for use in the production of PLD, we employed a series of combinatorial strategies to increase PLD expression in Bacillus subtilis WB600. These strategies included screening of signal peptides, selection of different plasmids, and optimization of the sequences of the ribosome-binding site (RBS) and the spacer region. We found that using the signal peptide amyE results in the highest extracellular PLD activity (11.3 U/ml) and in a PLD expression level 5.27-fold higher than when the endogenous signal peptide is used. Furthermore, the strain harboring the recombinant expression plasmid pMA0911-PLD-amyE-his produced PLD with activity enhanced by 69.03% (19.1 U/ml). We then used the online tool \RBS Calculator v2.0 to optimize the sequences of the RBS and the spacer. Using the optimized sequences resulted in an increase in the enzyme activity by about 26.7% (24.2 U/ml). In addition, we found through a transfer experiment that the retention rate of the recombinant plasmid after 5 generations was still 100%. The final product, phosphatidylserine (PS), was successfully detected, with transphosphatidylation selectivity at 74.6%. This is similar to the values for the original producer.

The atomization characteristics of the surrogate fuel of diesel from indirect coal liquefaction at engine conditions

( Zhong Huang ),( Tingting Zhang ),( Dehao Ju ),( Xinqi Qiao ),( Jin Xiao ),( Zhen Huang ) 한국액체미립화학회 2017 한국액체미립화학회 학술강연회 논문집 Vol.2017 No.-

The fuel spray behavior has a significant impact on the performance and emission characteristics of the compression ignition engine. It controls the fuel air mixing, which in turns is critical for the combustion process. The differences in the physical properties of convention diesel and the diesel from coal indirect liquefaction (DICL) are expected to result in differences in their spray characteristics, thus, in the performance and emission characteristics of the engine. The aim of this work is to analyze the atomization characteristics of the DICL and compare the global and local flow characteristics of the DICL with the NO.2 diesel (D2) at engine conditions. A surrogate fuel of the DICL (a mixture of 72.4% n-dodecane and 27.6% methylcyclohexane by mass) was built according to its components to simulate the atomization characteristics of the DICL in the non-reacting high temperature and high pressure environment. The implicit Large Eddy Simulation (LES) was used together with the Lagrangian particle tracking approach for the liquid phase in the present study. The simulation results show that the liquid penetration of the DICL is shorter than that of theD2, while there is no significant difference in the vapor penetration of the two fuels. In addition, the gas-phase axial velocity of the DICL along the spray center line is slightly larger than that of D2 in upstream of the spray. Moreover, the SMD of the DICL is larger than that of D2. The studies on the effects of the injection pressure on the atomization characteristics of the DICL were also carried out. The results imply that the injection pressure of the DICL is not required as large as the injection pressure for D2 to generate the same spray quality. This study is helpful to understand the differences in the atomization characteristics between DICL and D2.

다중완화시간 격자 볼츠만기법을 이용한 다양한 종횡비의 리드드리븐 공동유동 수치해석연구

황팅팅(Tingting Huang),송주헌(Juhun Song),임희창(Hee-Chang Lim) 한국가시화정보학회 2020 한국가시화정보학회지 Vol.18 No.3

This study performs a numerical simulation of lid driven rectangular cavity flow with different aspect ratios of k = 0.5 to 4 under Reynolds 100, 1,000, 10,000 by using multi-relaxation time (MRT) Lattice Boltzmann Method (LBM). In order to achieve better convergence, well-posed boundary conditions in the domain should be defined such as no-slip condition on side and bottom solid wall surfaces and uniform horizontal velocity on the top of the cavity. This study focuses on the flow inside different shape of rectangular cavity with the aim to observe the effect of the Reynolds number and aspect ratio on the flow characteristics and primary/secondary vortex formation. In order to validate the study, the results have been compared with existing works. The result shows that the Reynolds number and the aspect ratio both has substantial effects on the flow inside the lid-driven rectangular cavity.

A study on the Influencing Factors to China-Pakistan Trade under BRI Initiative

Jun Tang(Jun Tang),Xuanyu Xing(Xuanyu Xing),Tingting Huang(Tingting Huang) 아시아사회과학학회 2023 Jornal of Asia Social Science Vol.10 No.1

The Belt and Road Initiative is a grand plan proposed by China to promote the common development and prosperity of China and its partner countries. With the construction of the China-Pakistan Economic Corridor, the flagship of the Belt and Road Initiative, China-Pakistan relations are getting closer, known as the “iron brother”. Economic cooperation is a key priority for both sides. Under the context of the Belt and Road Initiative, this study uses trade data of China and Pakistan to find out the bilateral trade problems and the influencing factors behind them. This study will discuss the above questions and propose some suggestions for policymakers. Specifically, the discussion of the problems was conducted in terms of GDP, population, comparative advantages, international environment, domestic environment, and other factors respectively. In the conclusion part, the author suggests that the two countries should try to improve their domestic/international environment and reduce policy barriers to promote the trade. Modern industrial parks are encouraged to solve the problems. This study on the influencing factors and economic trade relations between China and Pakistan hopes to provide some academic references to the study of the CPEC and provide some guidance to the policy makers.

Adaptive Event-triggered Control of a Class of Series Elastic Actuator System

Tingting Gao,Jiangshuai Huang,Rui Ling,Yong Zhou 제어·로봇·시스템학회 2021 International Journal of Control, Automation, and Vol.19 No.7

In this paper, the event-triggered adaptive control for a class of series elastic actuator systems is considered. To solve this problem, firstly we propose an adaptive control scheme with a novel one-step design framework such that both the controller expression and parameter estimator are much simpler than related existing recursive design approaches. Then a set of event-triggering conditions is designed which is updated for each triggering. The ISS assumption is not needed for control design. It is shown that the proposed control schemes guarantee that allthe closed-loop signals are semi-globally bounded and the stabilization error converges to the origin asymptotically. The Zeno behavior is avoided. Simulation results illustrate the effectiveness of our scheme.

Major ginsenosides from Panax ginseng promote aerobic cellular respiration and SIRT1-mediated mitochondrial biosynthesis in cardiomyocytes and neurons

Huang, Qingxia,Lou, Tingting,Lu, Jing,Wang, Manying,Chen, Xuenan,Xue, Linyuan,Tang, Xiaolei,Qi, Wenxiu,Zhang, Zepeng,Su, Hang,Jin, Wenqi,Jing, Chenxu,Zhao, Daqing,Sun, Liwei,Li, Xiangyan The Korean Society of Ginseng 2022 Journal of Ginseng Research Vol.46 No.6

Background: Aerobic cellular respiration provides chemical energy, adenosine triphosphate (ATP), to maintain multiple cellular functions. Sirtuin 1 (SIRT1) can deacetylate peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α) to promote mitochondrial biosynthesis. Targeting energy metabolism is a potential strategy for the prevention and treatment of various diseases, such as cardiac and neurological disorders. Ginsenosides, one of the major bioactive constituents of Panax ginseng, have been extensively used due to their diverse beneficial effects on healthy subjects and patients with different diseases. However, the underlying molecular mechanisms of total ginsenosides (GS) on energy metabolism remain unclear. Methods: In this study, oxygen consumption rate, ATP production, mitochondrial biosynthesis, glucose metabolism, and SIRT1-PGC-1α pathways in untreated and GS-treated different cells, fly, and mouse models were investigated. Results: GS pretreatment enhanced mitochondrial respiration capacity and ATP production in aerobic respiration-dominated cardiomyocytes and neurons, and promoted tricarboxylic acid metabolism in cardiomyocytes. Moreover, GS clearly enhanced NAD⁺-dependent SIRT1 activation to increase mitochondrial biosynthesis in cardiomyocytes and neurons, which was completely abrogated by nicotinamide. Importantly, ginsenoside monomers, such as Rg1, Re, Rf, Rb1, Rc, Rh1, Rb2, and Rb3, were found to activate SIRT1 and promote energy metabolism. Conclusion: This study may provide new insights into the extensive application of ginseng for cardiac and neurological protection in healthy subjects and patients.

Identification and Characterization of a New Alkaline SGNH Hydrolase from a Thermophilic Bacterium Bacillus sp. K91

( Tingting Yu ),( Junmei Ding ),( Qingxia Zheng ),( Nanyu Han ),( Jialin Yu ),( Yunjuan Yang ),( Junjun Li ),( Yuelin Mu ),( Qian Wu ),( Zunxi Huang ) 한국미생물 · 생명공학회 2016 Journal of microbiology and biotechnology Vol.26 No.4

est19 is a gene from Bacillus sp. K91 that encodes a new esterase. A comparison of the amino acid sequence showed that Est19 has typical Ser-Gly-Asn-His (SGNH) family motifs and could be grouped into the SGNH hydrolase family. The Est19 protein was functionally cloned, and expressed and purified from Escherichia coli BL21(DE3). The enzyme activity was optimal at 60°C and pH 9.0, and displayed esterase activity towards esters with short-chain acyl esters (C2-C6). A structural model of Est19 was constructed using phospholipase A1 from Streptomyces albidoflavus NA297 as a template. The structure showed an α/β-hydrolase fold and indicated the presence of the typical catalytic triad Ser49-Asp227-His230, which were further investigated by site-directed mutagenesis. To the best of our knowledge, Est19 is a new member of the SGNH hydrolase family identified from thermophiles, which may be applicable in the industrial production of semisynthetic β-lactam antibiotics after modification.

Comparison of Triangular Silver Nanoprisms with Different Capping Agents and Structural Size for H2O2 Etching-Based Biosensors

Lishi Huang,Caihong Yuan,Wenli Chen,Fanshu Zeng,Hui Xu,Yahui Zhang,Tingting Jiang 성균관대학교(자연과학캠퍼스) 성균나노과학기술원 2018 NANO Vol.13 No.2

This study compared the susceptibility of different triangular silver nanoprisms (TSNPRs) towards the etching of hydrogen peroxide (H2O2), a catalytical product of glucose oxidase (GOx). The influence of capping agents and structural size have been explored towards the oxidation of silver nanoprisms. Results indicated that the etching of the TSNPRs was extremely effected by surface capping agents, in which citrate contributed a highest H2O2-sensitive effect in the absence of secondary capping ligands (e.g., glycerol and ethanol). Meanwhile, compared to bigger TSNPRs, smaller nanoprisms exhibited a different signal output of plasma resonance peak through intensity decrease rather than wavelength shift, making them more H2O2-etching susceptibile. In virtue of GOx etching-based system, TSNPRs with a small size and citrate capping were served as a substitute for big nanoprisms to sense glucose, offering a number of advantages such as high sensitivity, improved calibration, time-saving and extended detection ranges. Moreover, the small sized TSNPRs capping with citrate alone have been expected to be of great interest in the trace of GOx, providing an ultrahigh sensitive GOx etching-based analytical platform for point-of-care diagnostics towards other analytes (e.g., DNA, protein).

Dual-Responsive Cross-Linked Micelles from Amphiphilic Four-Arm Star Copolymers with Different Block Ratios for Triggering DOX Release

Yunwei Huang,Yanzhe Li,Zilun Tang,Qiuping Su,Tingting Liao,Yuxin Gu,Xiaofeng Lin,Xihong Zu,Wenjing Lin,Guobin Yi 한국고분자학회 2020 Macromolecular Research Vol.28 No.8

The four-arm star copolymers poly(methacrylic acid)-poly(2-hydroxyethyl methacrylate-disulfide~)-poly(poly(ethylene glycol) methyl ether methacrylate) (4AS-PMAAx-(PHEMA-SS~)y-PPEGMAz) with four different block ratios were synthesized and could self-assembled into cross-linked polymer micelles for the exploration of the structure-property relationship. The cross-linked polymer micelles in aqueous solution had low critical micelle concentration (CMC) values (1.9-4.6 mg/L), which exhibited better stability than non-cross-linked micelles. The CMC value decreased with the increase of the length of inner PMAA core and hydrophobic PHEMA cross-linked middle layer. The blank and doxorubicin (DOX)- loaded micelles with different block ratios were prepared by dialysis with the particle sizes of 120-240 nm. The longer inner PMAA core and cross-linked middle layer enhanced the drug loading content (DLC) results and led to relatively bigger particle sizes of polymer micelles. The in vitro DOX release data revealed that DOX-loaded micelles had low DOX cumulative release percentages of 18-37% after 110 h at pH 7.4, but up to 83-90% when introducing reductant GSH at pH 5.0. The 4AS-PMAA21.2-(PHEMASS~) 13.1-PPEGMA5.1 micelles with the longest PMAA core had the largest cumulative release of 90.1%. The DOX release process and mechanism of the micelles at different conditions fitted well with the semi-empirical equation. Overall, the results demonstrated that the block ratios and pH/redox-responsiveness of these four-arm star copolymers could be well-controlled and their self-assembled cross-linked micelles as anticancer drug carrier system could be improved by optimizing the different ratios.

Zearalenone exposure affects the Wnt/β-catenin signaling pathway and related genes of porcine endometrial epithelial cells in vitro

Song Tingting,Yang Weiren,Huang Libo,Yang Zaibin,Jiang Shuzhen 아세아·태평양축산학회 2021 Animal Bioscience Vol.34 No.6

Objective: Zearalenone (ZEA) has estrogen-like effects. Our previous study has shown that ZEA (0.5 to 1.5 mg/kg) could induce abnormal uterine proliferation through transforming growth factor signaling pathway. To further study the other regulatory networks of uterine hypertrophy caused by ZEA, the potential mechanism of ZEA on porcine endometrial epithelial cells (PECs) was explored by the Illumina Hiseq 2000 sequencing system. Methods: The PECs were treated with ZEA at 0 (ZEA0), 5 (ZEA5), 20 (ZEA20), and 80 (ZEA80) μmol/L for 24 h. The collected cells were subjected to cell cycle, RNA-seq, real-time quantitative polymerase chain reaction, immunofluorescence, and western blot analysis. Results: The proportion of cells in the S and G2 phases decreased (p<0.05), but the proportion of cells in the G1 phase increased (p<0.05) in the ZEA80 treatment. Data analysis revealed that the expression of Wnt pathway-related genes, estrogen-related genes, and mitogen-activated protein kinase pathway-related genes increased (p<0.05), but the expression of genetic stability genes decreased (p<0.05) with increasing ZEA concentrations. The relative mRNA and protein expression of WNT1, β-catenin, glycogen synthase kinase 3β (GSK-3β) were increased (p<0.05) with ZEA increasing, while the relative mRNA and protein expression of cyclin D1 (CCND1) was decreased (p<0.05). Moreover, our immunofluorescence results indicate that β-catenin accumulated around the nucleus from the cell membrane and cytoplasm with increasing ZEA concentrations. Conclusion: In summary, ZEA can activate the Wnt/β-catenin signaling pathway by upregulating WNT1 and β-catenin expression, to promote the proliferation and development of PECs. At the same time, the up-regulation of GSK-3β and down-regulation of CCND1, as well as the mRNA expression of other pathway related genes indicated that other potential effects of ZEA on the uterine development need further study. Objective: Zearalenone (ZEA) has estrogen-like effects. Our previous study has shown that ZEA (0.5 to 1.5 mg/kg) could induce abnormal uterine proliferation through transforming growth factor signaling pathway. To further study the other regulatory networks of uterine hypertrophy caused by ZEA, the potential mechanism of ZEA on porcine endometrial epithelial cells (PECs) was explored by the Illumina Hiseq 2000 sequencing system.Methods: The PECs were treated with ZEA at 0 (ZEA0), 5 (ZEA5), 20 (ZEA20), and 80 (ZEA80) μmol/L for 24 h. The collected cells were subjected to cell cycle, RNA-seq, real-time quantitative polymerase chain reaction, immunofluorescence, and western blot analysis.Results: The proportion of cells in the S and G2 phases decreased (p<0.05), but the proportion of cells in the G1 phase increased (p<0.05) in the ZEA80 treatment. Data analysis revealed that the expression of Wnt pathway-related genes, estrogen-related genes, and mitogen-activated protein kinase pathway-related genes increased (p<0.05), but the expression of genetic stability genes decreased (p<0.05) with increasing ZEA concentrations. The relative mRNA and protein expression of WNT1, β-catenin, glycogen synthase kinase 3β (GSK-3β) were increased (p<0.05) with ZEA increasing, while the relative mRNA and protein expression of cyclin D1 (CCND1) was decreased (p<0.05). Moreover, our immunofluorescence results indicate that β-catenin accumulated around the nucleus from the cell membrane and cytoplasm with increasing ZEA concentrations.Conclusion: In summary, ZEA can activate the Wnt/β-catenin signaling pathway by up-regulating WNT1 and β-catenin expression, to promote the proliferation and development of PECs. At the same time, the up-regulation of GSK-3β and down-regulation of CCND1, as well as the mRNA expression of other pathway related genes indicated that other potential effects of ZEA on the uterine development need further study.