Apocynaceae系细胞尿苷二磷酸葡萄糖焦磷酸化酶的纯化和表征

经4步分离纯化的Apocynaceae植物培养细胞尿苷二磷酸葡萄糖焦磷酸化酶(UGPase)的比活为2049Umg,与细胞粗提液相比,活力提高97倍,活力回收率为21%,可直接用于尿苷二磷酸葡萄糖(UDPG)的合成,并且储存稳定性很好。UGPase的最适pH值为72,UGPase的最适温度为37℃左右。Mg²⁺是UGPase发挥活力所必需的。高浓度UTP和Glc6P均能抑制UGPase的活力。     

p38 MAPK Inhibitor (SB203580) and Metformin Reduces Aortic Protein Carbonyl and Inflammation in Non-obese Type 2 Diabetic Rats

Microvascular and macrovascular diseases are the main causes of morbidity in type 2 diabetes patients through chronic hyperglycaemic condition via oxidative stress and inflammation. Reactive oxygen species (ROS) activate p38 MAPK phosphorylation and inflammation which enhances protein modification by carbonylation. The use of metformin and a p38 MAPK inhibitor is hypothesised to reduce ROS production and inflammation but effects of metformin and p38 MAPK inhibitor (SB203580) on ROS production and inflammation in vascular type 2 diabetes mellitus non-obese (T2DM) have not been investigated. The Goto-Kakizaki rat T2DM model was divided into three groups as T2DM, T2DM treated with 15 mg/kg bw metformin and T2DM treated with 2 mg/kg bw SB203580 for 4 weeks. Rat aortas were isolated and protein carbonyl (PC) contents were measured by spectrophotometric DNPH assay. Aortic IL-1ß level was determined by ELISA. Results showed that aortic PC contents in the T2DM group were significantly higher than in non-diabetic rats. Treatment with metformin or SB203580 significantly reduced PC contents while only metformin significantly reduced IL-1ß levels. Findings indicated that metformin reduced ROS production and inflammation in diabetic vessels and possibly reduce vascular complications in non-obese T2DM.     

Biochemical characterization of the recombinant schistosome tegumental protein SmALDH_312 produced in E. coli and baculovirus expression vector system

BackgroundThe heterologous expression of parasitic proteins is challenging because the sequence composition often differs significantly from host preferences. However, the production of such proteins is important because they are potential drug targets and can be screened for interactions with new lead compounds. Here we compared two expression systems for the production of an active recombinant aldehyde dehydrogenase (SmALDH_312) from Schistosoma mansoni, which causes the neglected tropical disease schistosomiasis.ResultsWe produced SmALDH_312 successfully in the bacterium Escherichia coli and in the baculovirus expression vector system (BEVS). Both versions of the recombinant protein were found to be active in vitro, but the BEVS-derived enzyme showed 3.7-fold higher specific activity and was selected for further characterization. We investigated the influence of Mg²⁺, Ca²⁺ and Mn²⁺, and found out that the specific activity of the enzyme increased 1.5-fold in the presence of 0.5 mM Mg²⁺. Finally, we characterized the kinetic properties of the enzyme using a design-of-experiment approach, revealing optimal activity at pH 7.6 and 41°C.ConclusionsAlthough, E. coli has many advantages, such as rapid expression, high yields and low costs, this system was outperformed by BEVS for the production of a schistosome ALDH. BEVS therefore provides an opportunity for the expression and subsequent evaluation of schistosome enzymes as drug targets.How to cite: Harnischfeger J, Beutler M, Salzig D, et al. Biochemical characterization of the recombinant schistosome tegumental protein SmALDH_312 produced in E. coli and baculovirus expression vector system. Electron J Biotechnol 2021;54. https://doi.org/10.1016/j.ejbt.2021.08.002     

串联亲和纯化技术筛选hCLP46的相互作用蛋白

hCLP46(human CAP10-like protein46)是从MDS-AML患者的CD34+干细胞cDNA文库中筛选出的基因.我们利用串联亲和纯化技术来筛选与hCLP46有相互作用的蛋白.通过体内交联-甘氨酸洗脱策略,检测到7条有差异的蛋白带,经液相色谱-质谱联用鉴定,得到了CNX和PDI等一系列内质网伴侣蛋白.所以hCLP46可能是一个糖蛋白,其成熟过程利用了BiP/Grp94和CNX/CRT 2套伴侣蛋白系统.     

医学人文教育创新创业实践初探

该文主要介绍在应用型本科医学院校中,医学人文教育借助大学生创新创业项目孵化,申报实用新型专利,实践人文、环保、科技的创新理念,实现医学人文教育的创新突破,将人文关怀渗透到人们的生活环境中,助力健康中国建设。     

铜胁迫对小叶黄杨生理生态特征的影响

采用水培试验的方法,研究不同浓度铜（0、20、40、60、80和100mg/L）胁迫条件下小叶黄杨的生长和生理特征。结果表明,随铜浓度的增加,小叶黄杨的生物量（鲜重）逐渐下降;蛋白质含量呈下降趋势;叶绿素含量先上升后下降;丙二醛的含量逐渐增加;超氧化物歧化酶和过氧化物酶的活性均呈先增加后下降的趋势。     

活化AMPK促进骨骼肌蛋白质降解过程中FoxO3 mRNA的表达

目的通过给大鼠注射AMPK激动剂AICAR,提高大鼠骨骼肌中AMPK的活性,观察FoxO3 mRNA表达量的变化,探讨骨骼肌蛋白质的降解机制。方法 24只雄性SD大鼠,8月龄,180～200 g,根据用药后的取材时间分4组:AICAR注射后1 h组、2 h组、7 h组,生理盐水对照组。采用同位素技术测定腓肠肌中AMPK活性变化,采用荧光定量PCR技术,测定腓肠肌中MuRF1、MAFbx、FoxO3基因表达量的变化。结果 AICAR注射后1 h组、2 h组、7 h组,AMPK活性与对照组相比显著性升高(P<0.05或P<0.01),AICAR注射后7 h组,AMPK活性开始下降;AICAR注射后1 h组、2 h组,MAFbx mRNA、MuRF1mRNA表达量与对照组相比非常显著性升高(P<0.01),分别升高2.45、2.23倍和2.67、2.30倍;AICAR注射后7 h组,MuRF1 mRNA表达量显著性升高(P<0.05),升高2.01倍;FoxO3mRNA表达量在AICAR注药组与对照组之间没有显著差异。结论 研究认为大鼠一次性注射AICAR,提高AMPK活性,可能不是通过促进FoxO3 mRNA的表达,促进骨骼肌蛋白质的降解。     

支链氨基酸与运动的关系

本文在广泛查阅相关文献的基础上,就支链氨基酸与能量代谢、蛋白质代谢,支链氨基酸与运动性中枢疲劳、外周疲劳以及支链氨基酸的毒性等方面加以综述并进行可行性分析论证。结果表明:支链氨基酸与人体运动能力关系密切,能够为长时间的耐力运动提供能量储备,促进肌肉蛋白质的合成,而且可以调节引发运动性疲劳的外周机制,推迟运动性疲劳的出现,运动后补充支链氨基酸还可以促进运动性疲劳的恢复。但至于支链氨基酸对蛋白质分解代谢的影响及其抗中枢疲劳效应,在长期训练中补充BCAA是否有减少和修复肌肉损伤的能力,是否存在性别差异以及服用大量BCAA是否对大脑和肌肉产生消极作用,导致肠胃功能的紊乱则还有待进一步的研究与探索。     

植物病毒抗血清的制备及IgG纯化与酶标技术研究

以烟草花叶病毒（TMv）为例研究了制备植物病毒抗血清的一般方法．以家兔和小白鼠为免疫动物，运用饱和硫酸接沉淀法制备出高效价的兔和瓦抗TMV血清，并运用先进的ELISA方法测定了抗血清中抗体IgG的效价；用醋酸纤维凝胶电泳技术分析了抗血清中抗体的lgG纯度，应用免疫双扩散法检测了提纯的IgG的生物活性．同时还运用成二醛一步法制出高结合率的IgG与过氧化氢酶（HRP）标记的抗体，形成了一整套制备抗血清、纯化IgG并对IgG进行酶标的技术，为植物病毒检测提供了快速简便的方法．     

云烟85种子萌发过程物质消耗研究

以烤烟品种云烟85种子为实验材料,从种子吸水前0h和吸水后的24h,48h,72h,96h,120h,144h,168h分8个时期分析了烟草种子萌发时各种养分的动态变化。结果表明:种子萌发至168h时,种子内可溶性总糖和还原糖含量达最大值,随后可溶性总糖和还原糖含量均维持在一个稳定的水平;种子萌发至24h时,种子内的蛋白质含量开始下降,至168h时达到最低;种子萌发至72h时,种子内的粗脂肪含量开始下降,至168h时达到最低。