用AFM研究杜仲抗真菌蛋白的晶体生长

随着后基因组时代的到来以及蛋白质组学研究的深入开展,研究蛋白质晶体生长成为生物化学和结构生物学领域一个广泛关注的课题.通过使用原子力显微镜(Atomic Force Microscope,简称AFM)对杜仲抗真菌蛋白(eucommia antifungal protein,简称EAFP)的晶体在有母液存在下原位实时动态地进行了晶面生长观察.研究结果表明:不同过饱和度对EAFP晶体生长形貌的影响较大,较高的过饱和度下生长很快,生长台阶密度高,较高的过饱和度下主要进行各向异性二维台阶的发生、发展,较低的过饱和度下主要采用螺旋位错的生长方式,当过饱和度极低时生长缓慢,且晶体表面有很多小孔存在,晶面生长很不完整;还对不同过饱和度下晶体生长速率进行了定量的测量,也反映了过饱和度对EAFP晶体生长的影响;同时对在AFM观察过程中由探针的扫描速度和方向对表面形貌的影响进行了讨论.     

天花粉蛋白N－糖苷酶的活性

本文报道了由栝楼根制备的天花粉蛋白具有良好的引产活性和对免疫系统有一定的抑制作用，其本质是天花粉蛋白具有Ｎ－糖苷酶的活性。     

天花粉蛋白单克隆抗体杂交瘤细胞系的制备及鉴定

将小鼠骨髓瘤细胞（SP2／0细胞）与经TCS免疫过的小鼠脾细胞融合,融合后的细胞悬浮于HAT培养液中进行选择性培养,挑选可以分泌TCS单克隆抗体的细胞系,再经克隆化为单克隆细胞系。用间接酶联免疫吸附法检测此细胞系分泌单克隆抗体情况。经细胞融合和2次克隆化后,获得1株可以稳定分泌TCS单克隆抗体杂交瘤细胞系,ELISA检测结果为阳性。获得了可以稳定分泌天花粉蛋白单克隆抗体的杂交瘤细胞系,为TCS鉴定、应用奠定良好的基础。     

原子力显微镜研究聚四氟乙烯纳米粒子对瑞氏木霉疏水蛋白HFBII的吸附

利用离子束溅射镀膜技术将聚四氟乙烯(Teflon)纳米粒子沉积到石英基板,将基板置于50 μg/mL的瑞氏木霉疏水蛋白HFBII溶液中浸泡,通过原子力显微镜直接观察和水接触角测试仪测试基板表面水接触角的变化,研究了HFBII在Teflon纳米粒子表面的吸附行为.结果表明Teflon纳米粒子能够对HFBII产生吸附且对其生长有很强的异相成核作用,同时用超声波方法对Teflon纳米粒子上的HFBII进行脱附,并通过原子力显微镜图像进行了分析.     

原子力显微镜研究聚四氟乙烯纳米粒子对瑞氏木霉疏水蛋白HFBⅡ的吸附

利用离子束溅射镀膜技术将聚四氟乙烯（Teflon）纳米粒子沉积到石英基板,将基板置于50μg/mL的瑞氏木霉疏水蛋白HFBⅡ溶液中浸泡,通过原子力显微镜直接观察和水接触角测试仪测试基板表面水接触角的变化,研究了HFBⅡ在Teflon纳米粒子表面的吸附行为．结果表明Teflon纳米粒子能够对HFBⅡ产生吸附且对其生长有很强的异相成核作用,同时用超声波方法对Teflon纳米粒子上的HFBⅡ进行脱附,并通过原子力显微镜图像进行了分析．     

天花粉蛋白结构改造对引产活性的影响

目的：研究结构改造对天花粉蛋白（ＴＣＳ）引产活性的影响。方法：利用定点突变将ＴＣＳ第２１９位的天冬酰胺残基改变为半胱氨酸残基（Ｑ２１９Ｃ）以供定点聚乙二醇（ＰＥＧ）修饰,测定修饰产物的致本失活活性及小鼠中期引产活性。结果：ＰＥＧ修饰使的致核糖体失活活性约降低９０％;相对分子质量为５０００的ＰＥＧ修饰对引产活性无明显影响,而相对分子质量为２００００的ＰＥＧ修饰却明显增强引产活性;此外,ＰＥＧ修饰使Ｔ     

天花粉蛋白抗单纯疱疹病毒1型的实验研究

检测用基因工程方法制备的天花粉蛋白抗单纯疱疹病毒1型的活性。将单纯疱疹病毒1型在细胞系中培养．加入天花粉蛋白后,观察细胞病变情况,用酶联免疫吸附（ELISA）的方法检测病毒生长情况。结果显示,加入天花粉蛋白的细胞始终未见细胞病变,ELISA检测为阴性;阴性对照组细胞在第3d以后均出现单纯疱疹病毒1型感染典型细胞病变．ELISA检测结果为阳性;阳性对照组始终未见单纯疱疹病毒1型感染典型细胞病变,ELISA检测结果为阴性。天花粉蛋白在体外培养细胞系中可以抑制单纯疱疹病毒1型。     

杜仲抗真菌蛋白(EAFP)晶体动态生长研究

杜仲抗真菌蛋白(eucommia antifungal protcin,EAFP),含有41个氨基酸残基,其中有10个半胱氨酸,半胱氨酸间形成的二硫桥键使分子构象很稳定.EAFP的晶体属单斜晶系,空间群为P2 1.晶胞参数为:a=1.908 5 nm,b=2.322 5 nm,c=3.085 4 nm10-6,β=98.64,分子量为4158.9.EAFP晶体生长速度快,对X射线的衍射能力强,很值得研究其生长的机理.利用原子力,显微镜(AFM)对EAFP晶体的{100}面进行动态的生长研究,发现在中低过饱和度下主要以各向异性的单双链螺旋位错的生长模式进行生长,详细研究了这种螺旋的形成机制,并探讨了其结构基础.     

原子力显微镜研究蛋白质晶体生长机理及进展

原子力显微镜(AFM)这种能进行微尺度测量的精密仪器是目前研究晶体生长机理最为有效的方法之一,在晶体生长机理研究中起到了重要的作用,尤其是对于蛋白质晶体生长的研究显得更为突出.分别简明地阐述了利用AFM进行蛋白质晶体生长研究的工作原理,以及在这方面已经取得的研究成果,包括二维台阶生长、螺旋位错生长、法向生长、三维成核生长、台阶发展的不对称性、及杜仲抗真菌蛋白(EAFP)和天花粉蛋白(TCS)晶体生长的新进展.论述了利用空间微重力环境进行蛋白质晶体生长及空间蛋白质晶体生长的科研和应用, 指出今后进行蛋白质晶体生长研究的趋势和前景.     

Research progress in quantifying the mechanical properties of single living cells using atomic force microscopy

The advent of atomic force microscopy （AFM） provides a powerful tool for investigating the behaviors of single living cells under near physiological conditions. Besides acquiring the images of cellular ultra-microstruc- tures with nanometer resolution, the most remarkable advances are achieved on the use of AFM indenting tech- nique to quantify the mechanical properties of single living ceils. By indenting single living cells with AFM tip, we can obtain the mechanical properties of cells and monitor their dynamic changes during the biological processes （e.g., after the stimulation of drugs）. AFM indentation-based mechan- ical analysis of single cells provides a novel approach to characterize the behaviors of cells from the perspective of biomechanics, considerably complementing the traditional biological experimental methods. Now, AFM indentation technique has been widely used in the life sciences, yielding a large amount of novel information that is meaningful to our understanding of the underlying mechanisms that govern the cellular biological functions. Here, based on the authors＇ own researches on AFM measurement of cellular mechani- cal properties, the principle and method of AFM indentationtechnique was presented, the recent progress of measuring the cellular mechanical properties using AFM was summa- rized, and the challenges of AFM single-cell nanomechani- cal analysis were discussed.     

用AFM观测凝胶法下KClO4晶体的生长机理

用原子力显微镜(atomic force microscopy,AFM)对凝胶法生长的KClO4晶体的最重要面{001}面微观形貌进行了研究。结果表明,在不同的过饱和度下,KClO4晶体分别表现为二维成核生长、三维成核生长、层核生长以及多层堆垛生长等生长机理。从热力学角度解释了过饱和度对KClO4晶体生长机理的影响。对二维成核生长机理,揭示了KClO4晶体生长中二维核的形状和取向,及生长台阶的取向。研究结果还表明,杂质的存在将阻止台阶的向前推进,并导致聚并台阶及弯曲台阶的形成。     

The asymmetric membrane structure of erythrocytes from Crucian carp studied by atomic force microscopy

Cell membranes play a key role in cellular activities. Fish erythrocytes, prototype of the nucleated erythrocytes of lower vertebrates, are predominantly oval, biconvex discs with elliptical nucleus and much larger in size than human erythrocytes. In attempts to disclose the correlation of membrane structure between fish and human erythrocytes, we used in situ atomic force microscope （AFM） combined with single-molecule force spectroscopy to study the membrane structure of Crucian carp erythrocytes under quasi-native conditions. Our results revealed the asymmetric distribution of proteins in Crucian carp erythrocyte membrane： the outer leaflet of membrane is rather smooth without any proteins, whereas the inner leaflet of membrane is very rough with dense proteins. The asymmetry of fish erythrocyte membrane structure fits well with the semi-mosaic model of human erythrocyte membrane structure. This similarity of membrane structure between human and fish erythrocytes extends the semimosaic model of erythrocytes membrane structure to a wider range of species.     

聚丙烯酸纳米微凝胶的表征

利用原子力显微镜（AFM）对聚丙烯酸纳米微凝胶的形态、结构进行了表征,单体浓度、交 联剂与单体摩尔比对聚丙烯酸微凝胶的粒径都有一定影响,在实验条件下均可得到粒径为20 ～60nm的微凝胶。加入夺氢型光引发剂BP-4进行二次聚合,可以得到大分子纳米凝胶引发剂,其粒径比一次聚合所得微凝胶增大10～20nm;二次聚合时加入N-异丙基丙烯酰胺（NIPAM）可得到50～90nm、表面包覆PNIPAM的纳米凝胶。     

不同形核机制下对动态再结晶过程模拟研究

为了研究形核机制对多晶金属材料热加工过程的影响,通过建立动态再结晶的二维元胞自动机模型,模拟了3种形核机制下动态再结晶组织的演化过程,得到了热加工过程中再结晶平均晶粒尺寸的变化规律及生长动力学特征.模拟结果表明,再结晶平均晶粒尺寸的变化规律由形核机制和生长方式共同决定,但稳态时的平均晶粒尺寸只与总的形核数目有关;生长动力学曲线均呈S形,其变化规律符合JMAK理论;Avrami指数的大小取决于晶核的空间分布.     

Resistance to rice blast (Pyricularia oryzae) caused by the expression of trichosanthin gene in transgenic rice plants transferred through agrobacterium method

The gene of trichosanthin has been transferred into rice plants through agrobacterium method. The single copy insertion and the expression of foreign gene have been proved in regenerated plants. In antifungal assay the degrees of rice blast (Pyricularia oryzae) infection of the transgenic plants expressing trichosanthin and expressingGUS gene as control have been evaluated. The differences such as the time of disease symptom observed, the number of infected plants and damaged leaves, the growth of infected plants of the two transgenic plants after being inoculated by rice blast (Pyricularia oryzae) are significant. The transgenic plants with trichosanthin gene grew faster than the plants withGUS gene, even when humidity environment was removed. The results show that the transgenic plants that expressed trichosanthin are able to delay the infection of rice blast compared with the plants as control. In addition, no damage caused by the expression of trichosanthin gene in transgenic plants has been observed.