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Thermostability of Subtilisin Nattokinase Obtained by Site-Directed Mutagenesis

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Wuhan University Journal of Natural Sciences 2014年第3期19卷 229-234页

作者： weng meizhi DENG Xiongwei WU Jieyuan ZOU Guolin Jiangxi Province Blood Center College of Life Sciences Wuhan University Hongdu Chinese Medical Hospital

To study the thermostability of Nattokinase（subtilisin NAT,NK）,three double mutant plasmids（pET-28a-NKG61C/S98C,pET-28a-NKT22C/S87C,pET-28a-NKS24C/S87C）were constructed by site-directed *** enzymes were detected using SDS-PAGE and disulfide bond formation was detected using Western blotting *** was tested by rates of inactivation at certain *** results showed that disulfide bond was not formed within two cysteines and the thermostability of three double mutants was not increased compared with the wild-type *** thermostability of NK performed in Ca2＋was stronger than in ethylenediaminetetraacetic acid（EDTA）.But when the temperature reached 62℃,the enzymes rapidly denatured and inactivated even in the presence of Ca2＋.Although the thermostability of mutants was not increased,this study shows a tendency of improving thermostability of NK in protein engineering.

关键词： nattokinase subtilisin thermostability

Contents Variation Analysis ofγ-Amino Butyric Acid in Semen sojae praeparatum Fermentation Using Online Pre-Column Derivatization-HPLC

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Wuhan University Journal of Natural Sciences 2021年第1期26卷 55-61页

作者： weng meizhi HE Jing REN Jiaxiu LIANG Yonghong CHEN Qingfeng SU Mingsheng WANG Liyuan XIE Xiaomei College of Traditional Chinese Medicine Jiangxi University of Traditional Chinese MedicineNanchang 330004JiangxiChina Institute of Translational Medicine Nanchang UniversityNanchang 330031JiangxiChina Key Laboratory of Modem Preparation ofTCM Ministry of EducationJiangxi University of Traditional Chinese MedicineNanchang 330004JiangxiChina

This paper reported the contents variation analysis ofγ-amino butyric acid(GABA)in Semen sojae praeparatum(SSP)which is a famous traditional Chinese *** performance liquid chromatography(HPLC)was used and GABA was derivatized by online pre-column derivatization with o-phthalaldehyde(OPA).To validate this method,the precision,stability,repeatability and recovery were *** the concentration range from 0.0125 to 0.400 mg/m L,the calibration curve for GABA was linear and the regression equation was obtained with correlation coefficient(R2)of *** high levels of GABA exist in SSP and the content changes of GABA at different time points during the fermenting process were *** the"yellow cladding"stage,GABA level was very low or even undetectable;the"secondary fermentation"stage witnessed a rapid increase of GABA content to 1.39-5.52 mg/g,which remained stable after 18 days of"secondary fermentation".This study demonstrated that GABA was generated at the"secondary fermentation"stage,revealing the significance and rationality of the"secondary fermentation"stage in the fermenting process of *** the other hand,it suggested the downside of taking soy isoflavones as the only measurement in existing quality assessment and optimization approach for the fermenting process of SSP.

关键词： Semen sojae praeparatum γ-amino butyric acid(GABA) "secondary fermentation"process online pre-column derivatization-high performance liquid chromatography

Liuwei Dihuang pill suppresses metastasis by regulating the wnt pathway and disrupting β-catenin/T cell factor interactions in a murine model of triple-negative breast cancer

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Journal of Traditional Chinese Medicine 2019年第6期39卷 826-832页

作者： Zheng Lixiang Zheng Qing Yu Zhipeng Wang Jian Ren Xiaoying Gong Yan Yang Xue Hunag Ping weng meizhi Liu Hongning Liu Haizhou School of Basic Medicine Jiangxi University of Traditional Chinese MedicineNanchang 330004China Jiangxi University of Traditional Chinese Medicine Nanchang 330004China Department of Gastroenterology the First Affiliated Hospital of Nanchang UniversityNanchang 330006China Teaching and Research Office of Internal Medicine of Traditional Chinese Medicine Department of MedicineJiangxi College of Traditional Chinese MedicineFuzhou 340000China School of Humanities Jiangxi University of Traditional Chinese MedicineNanchang 330004China

OBJECTIVE:To investigate if the Liuwei Dihuang pill(LWDHP)can inhibit metastasis to the liver and lungs in mice bearing triple-negative breast cancer(TNBC),and the molecular mechanism underpinning this ***:Ninety-nine TNBC bearing-mice were distributed randomly to five groups:control(Con),paclitaxel(PTX),low-dose LWDHP(LLP,2.3 g·kg^-1·d^-1),middle-dose LWDHP(MLP,4.6 g·kg^-1·d^-1)and high-dose LWDHP(HLP,9.2 g·kg^-1·d^-1).The LWDHP were administered(p.o.)to the agonal *** morphology of BC cells was observed by hematoxylin&eosin *** of axin-2,β-catenin,T cell factor(TCF),cyclin-D1 and vascular endothelial growth factor(VEGF)was detected by western blotting or immunofluorescence.β-catenin/TCF-1 interaction was measured using a co-immunoprecipitation ***:After LWDHP treatment,metastasis of BC cells to the lungs and liver was inhibited,expression of axin-2 was increased,expression of TCF-1,β-catenin,cyclin-D1 and VEGF was decreased,andβ-catenin/TCF-1 interaction was ***:The LWDHP could inhibit metastasis of BC cells to the liver and *** molecular mechanism underlying this action may be regulation of protein expression andβ-catenin/TCF-1 interactions in the Wnt pathway.

关键词： Triple negative breast neoplasms Neoplasm metastasis Beta catenin T cell transcription factor 1 Wnt signaling pathway Liuwei Dihuang pill

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天然纤溶酶的来源及生产方法研究进展

中国酿造 2024年第2期43卷 29-34页

作者：彭粹盈王盼盼邓雄伟谢小梅翁美芝江西中医药大学中医学院江西南昌330004 江西中医药大学南昌市洪都中医院江西南昌330008

全球每年有1790万人丧命于心血管疾病,占全球死亡人数的31%。而在各种心血管病中,血栓形成是最常见的一种,如何有效防治血栓是一个全球性的重大医学问题。在血栓防治中,纤溶酶潜力巨大。纤溶酶是一种蛋白水解酶,对血栓栓塞性疾病的防治... 详细信息

全球每年有1790万人丧命于心血管疾病,占全球死亡人数的31%。而在各种心血管病中,血栓形成是最常见的一种,如何有效防治血栓是一个全球性的重大医学问题。在血栓防治中,纤溶酶潜力巨大。纤溶酶是一种蛋白水解酶,对血栓栓塞性疾病的防治非常重要。该文综述了血栓形成及纤溶酶的作用机制,总结了纤溶酶的来源(海洋微生物、发酵食品微生物、植物和动物)和生产方法,并对纤溶酶生产方法作出总结和展望,以期为相关领域研究提供参考。

关键词：纤溶酶蛋白酶血栓溶栓机理纤溶酶原激活剂

等待肾移植患者体内预存HLA抗体特点及致敏因素分析

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中国输血杂志 2014年第4期27卷 383-386页

作者：翁美芝孙瑜吴红肖莉熊莉李国良江西省血液中心江西南昌330077

目的分析研究等待肾移植手术的患者体内预存群体反应性抗体(PRA)特点及其产生原因。方法采用酶联免疫吸附法(ELISA)检测1 618名等待肾移植患者PRA,根据患者原有的输血史、妊娠史和移植史,将其分为5组:未输血未妊娠组(n=804);单纯输血组(... 详细信息

目的分析研究等待肾移植手术的患者体内预存群体反应性抗体(PRA)特点及其产生原因。方法采用酶联免疫吸附法(ELISA)检测1 618名等待肾移植患者PRA,根据患者原有的输血史、妊娠史和移植史,将其分为5组:未输血未妊娠组(n=804);单纯输血组(n=330);单纯妊娠组(n=229);输血合并妊娠组(n=196);有移植史组(n=59)。结果 1 618名等待肾移植患者中,未输血未妊娠组、单纯输血组、单纯妊娠组、输血合并妊娠组和有移植史组PRA阳性率分别为5.97%、18.18%、22.71%、48.47%和76.27%。患者出现频率较高的HLAⅠ类抗体有抗-A2、-A23、-A66和-B7等,HLAⅡ类抗体有抗-DR52、-DR53、-DQ5和-DQ6等。有移植史组患者产生DR53和DR52抗体的频率较高,占HLAⅡ类抗体总数的19.32%。结论不同个体对相同致敏因素反应不同;在相同致敏因素存在的情况下,男性和女性患者PRA阳性率差异无统计学意义;输血史、妊娠史和移植史可协同提高致敏率;抗体产生的频率与抗原频率分布不相一致;HLAⅡ类抗体中,DR52和DR53抗体的产生与移植物失功存在一定关联性。

关键词：肾移植群体反应性抗体致敏因素人类白细胞抗原

“科学前沿进课堂”——高校生物化学特色教学初探

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生命的化学 2017年第6期37卷 1095-1099页

作者：舒青龙冯洁翁美芝江西中医药大学南昌330004

生物化学是高校许多专业的专业基础课,该课程原理繁多、概念抽象,如何让学生掌握其知识与技术,激发学生学习热情是教学中值得研究的问题。本文根据生命科学研究迅猛发展的特点,并结合自己的教学经验,详细介绍了"科学前沿进课堂"的特色... 详细信息

生物化学是高校许多专业的专业基础课,该课程原理繁多、概念抽象,如何让学生掌握其知识与技术,激发学生学习热情是教学中值得研究的问题。本文根据生命科学研究迅猛发展的特点,并结合自己的教学经验,详细介绍了"科学前沿进课堂"的特色教学法,包括该教学法的特点、素材选择和准备、该教学法的课堂实施及其教学效果等,旨在促进与同行的教学交流,共同提高高校生物化学的教学质量。

关键词：生物化学科学前沿高校教学

基于学生自主学习能力培养的教学模式在《细胞生物学》课程中的应用

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中国细胞生物学学报 2020年第8期42卷 1381-1386页

作者：翁美芝刘升长谢燕飞舒青龙彭淑红郑里翔江西中医药大学中医学院南昌330004

长久以来,《细胞生物学》课程的教学大多沿用传统教学方法,即主要以教师讲、学生听为主,此教学方法强调对知识的传承,但学生的自主学习能力得不到充分发挥。在新形势下,自主学习能力是竞争激烈的现代社会重要的生存能力,对当代大学生来... 详细信息

长久以来,《细胞生物学》课程的教学大多沿用传统教学方法,即主要以教师讲、学生听为主,此教学方法强调对知识的传承,但学生的自主学习能力得不到充分发挥。在新形势下,自主学习能力是竞争激烈的现代社会重要的生存能力,对当代大学生来说,自主学习能力的培养比知识的获取更为重要。针对《细胞生物学》现有教学模式的不足,该文以学生自主学习能力培养为着眼点,在教学过程中,采用问题讨论式教学结合思维导图教学的教学模式,改变一考定成绩的考核评价体系,初步探究了旨在培养学生自主学习能力的《细胞生物学》课程教学模式。实施效果表明,该教学模式达到了预期教学目标,有助于培养学生的自主学习能力。

关键词：细胞生物学自主学习思维导图教学模式

Robust interface and excellent as-built mechanical properties of Ti–6Al–4V fabricated through laser-aided additive manufacturing with powder and wire

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International Journal of Minerals,Metallurgy and Materials 2025年第1期32卷 154-168页

作者： Fei weng Guijun Bi Youxiang Chew Shang Sui Chaolin Tan Zhenglin Du Jinlong Su Fern Lan Ng Key Laboratory for Liquid-Solid Structural Evolution and Processing of Materials Ministry of EducationShandong Engineering&Technology Research Center for Superhard MaterialSchool of Materials Science and EngineeringShandong UniversityJinan 250061China Institute of Intelligent Manufacturing Guangdong Academy of SciencesGuangzhou 510095China Singapore Institute of Manufacturing Technology 5 Cleantech Loop636732Singapore

The feasibility of manufacturing Ti-6Al-4V samples through a combination of laser-aided additive manufacturing with powder(LAAM_(p))and wire(LAAM_(w))was explored.A process study was first conducted to successfully circumvent defects in Ti-6Al-4V deposits for LAAM_(p) and LAAM_(w),*** the optimized process parameters,robust interfaces were achieved between powder/wire deposits and the forged substrate,as well as between powder and wire *** characterization results revealed the epitaxial prior β grains in the deposited Ti-6Al-4V,wherein the powder deposit was dominated by a finerα′microstructure and the wire deposit was characterized by lamellar α *** mechanisms of microstructure formation and correlation with mechanical behavior were analyzed and *** mechanical properties of the interfacial samples can meet the requirements of the relevant Aerospace Material Specifications(AMS 6932)even without post heat *** fracture occurred within the interfacial area,further suggesting the robust *** findings of this study highlighted the feasibility of combining LAAM_(p) and LAAM_(w) in the direct manufacturing of Ti-6Al-4V parts in accordance with the required dimensional resolution and deposition rate,together with sound strength and ductility balance in the as-built condition.

关键词： laser-aided additive manufacturing powder deposition wire deposition interfacial characteristic mechanical behavior

淡豆豉中产γ-氨基丁酸微生物对产毒黄曲霉菌的拮抗作用和毒素合成关键基因mRNA表达的影响

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中草药 2024年第5期55卷 1510-1517页

作者：戴家齐贺婧朱晓蓝王立元龙凯周立分杨安金翁美芝谢小梅江西中医药大学江西南昌330004 南昌大学转化医学研究院江西南昌330031 江西中医药大学附属医院江西南昌330006

目的考察从淡豆豉Sojae Semen Praeparatum(SSP)炮制过程中分离的15株产γ-氨基丁酸微生物对产毒黄曲霉菌的拮抗能力并筛选出具有抑制产毒黄曲霉菌生长和降解黄曲霉毒素B1(aflatoxin B1,AFB1)的高效拮抗菌,探究高效拮抗菌发酵产物对黄... 详细信息

目的考察从淡豆豉Sojae Semen Praeparatum(SSP)炮制过程中分离的15株产γ-氨基丁酸微生物对产毒黄曲霉菌的拮抗能力并筛选出具有抑制产毒黄曲霉菌生长和降解黄曲霉毒素B1(aflatoxin B1,AFB1)的高效拮抗菌,探究高效拮抗菌发酵产物对黄曲霉毒素(aflatoxins)合成关键基因mRNA表达量的影响。方法运用平板对峙法与十字交叉法检测15株产γ-氨基丁酸微生物及其发酵产物对产毒黄曲霉标准株(简称:产毒标准株)生长的影响,超高效液相色谱-串联质谱法(ultra performance liquid chromatography mass spectrometry,UPLC-MS/MS)检测15株菌对AFB1的降解效果,考察其对产毒黄曲霉菌的拮抗能力并筛选出高效拮抗菌;针对高效拮抗菌发酵产物分别运用菌丝干重法检测其对产毒标准株菌丝生长的影响,实时荧光定量PCR(real-time quantitative polymerase chain reaction,RT-qPCR)检测其对黄曲霉毒素合成关键基因(aflR、aflD、aflM、aflP)mRNA表达的影响。结果15株菌及其发酵产物对产毒标准株的生长抑制率分别在15.88%~56.05%和25.74%~52.12%,对AFB1降解率在2.74%~57.87%,表明它们对产毒黄曲霉菌均有一定的拮抗作用,并筛选出具有高效拮抗作用的黑曲霉菌Aspergillus niger(JC2)和枯草芽孢杆菌Bacillus subtilis(Xd1)。当高效拮抗菌发酵产物加入量为2000µL时,JC2和Xd1对产毒标准株菌丝生长的抑制率分别为73.82%和63.34%,且能明显抑制黄曲霉毒素合成关键基因aflR、aflD、aflM、aflP的mRNA表达。结论淡豆豉炮制中存在既能产γ-氨基丁酸又能明显抑制产毒标准株生长和产毒的拮抗菌,其拮抗作用可能通过抑制产毒标准株菌丝生长和毒素合成关键基因的mRNA表达。

关键词：淡豆豉 γ-氨基丁酸产毒黄曲霉菌黄曲霉毒素拮抗作用高效拮抗菌实时荧光定量PCR