豆薯种子中一种具有抗植物病毒活性的蛋白质的初步晶体学研究

从豆科植物豆薯 (Pachyrrhizuserosus,Leguminosae)的种子中分离纯化了一种具有抗植物病毒活性的蛋白质 ,分子量约为 1 5kD。在 2 0 % (w/v)PEG60 0 0 ,0 .1mol/L磷酸盐缓冲液 (pH7.4)条件下得到了可供衍射用的单晶 ,在MarResearchIP上进行了衍射数据收集 ,其衍射能力达 2 .3 。经MarDenzo软件包处理 ,确定其空间群为P2 1 2 1 2 1 ,晶胞参数为 :a =2 8.80 8,b =32 .840 ,c=1 40 .397 。根据Matthews常数估算 ,每个不对称单元含 1个分子。     

天花粉蛋白2.6分辨率的晶体学修正

本文报道了用溶剂过滤法改善电子密度图,依据Collins提出的一级结构,重新建立天花粉蛋白分子模型。使用2.6分辨率衍射强度数据和约束参数最小二乘法,对处于晶体学不对称单元中两个天花粉蛋白分子(共3828个非氢原子)进行晶体学修正。修正结果,偏离因子R=0.223,键长根均方误差r.m.s=0.023,新的分子模型和以2F_0-F_o为系数的电子密度图符合甚好。     

冬凌草甲素乙酰衍生物的晶体结构

冬凌草甲素是从抗癌中草药冬凌草中提取的一种有效成分。它的乙酰衍生物的晶体结构和分子结构已由X—射线衍射法中直接法解出,R因子被修正到0.067。其晶体属单斜晶系,空问群为P2_1,晶胞参数是:a=8.923(3),b=6.551(2),c=20.102(3),β=99.77(2)°晶胞中含有两个C_(22)H_(30)O_7分子和两个乙醇(溶剂)分子。 晶体结构分析表明,分子中含有部分共轭的α—亚甲基环戊酮的五员环。其内角均分别地远小于各自碳原子SP~2或SP~3杂化所要求的数值,而含有较大的内应力。     

高效凝胶过滤色谱法分离测定豆薯种子蛋白

用高效凝胶蛋白往分离豆薯种子蛋白租提液,结合光电二极管阵列检测器对分离的蛋白峰进行紫外光谱扫描来确认蛋白的纯度,测定了3种蛋白的分子量,采用邻苯二甲醛（OPA）柱后衍生法测定了豆薯种子蛋白的氨基酸含量。     

天花粉蛋白分子的三维结构

本文报道了我们最近建立的单斜晶胞内一个不对称单元中两个天花粉蛋白分子的模型,指出蛋白分子结构可以划分为大小两个结构域以及二级结构的一些特点,描述了独立单元中以一种非对称方式相关联的两个蛋白分子的差异及其相互作用,并解释了重原子衍生物的结合位置。     

正交晶系天花粉蛋白2.4分辨率的晶体结构和分子结构

在酸性条件下(pH=5.4)培养出正交晶系天花粉蛋白晶体。用面探测器收集了X射线衍射强度数据。以C2空间群天花粉蛋白2.6分辨率的晶体结构中的分子模型作为已知结构模型,用分子置换法解出正交晶系P2_12_12_1空间群天花粉蛋白的晶体结构。所得到的初始结构模型先后用模拟退火法和立体化学制约的最小二乘修正技术对原子坐标及温度因子进行修正。最后在2.4分辨率范围内模型的β因子为0.197,键长的均方根误差为0.021,键角距离的均方根误差为0.081。分子结构模型与(2Fo—Fc)为系数的电子云密度图吻合得很好。键长键角合理。结构测定结果表明,P2_12_12_1;空间群天花粉蛋白的分子的空间结构与单斜晶系C2空间群中的分子结构甚为相似。     

A DETERMINATION OF THE THREE-DIMENSIONAL STRUCTURE OF TRICHOSANTHIN AT LOW RESOLUTION

Trichosanthin, extracted from a Chinese herb medicine, is a plant protein having a molec-ular weight of ca. 24,000 and containing 219 amino acid residues. The crystal is mono-clinic, belonging to the space group C2, with the following cell dimensions: a=75.64A, b=75.52A, c=88.85A β= 99.51°. Each asymmetric unit contains two such protein molecules. The UO_2Ac_2 and K_2Pt(NO_2)_4 derivatives of trichosanthin have been prepared by soakingin the respective solutions of high dilution. The joint probability of isomorphous replacementand anomalous scattering has been used to calculate the native phases. In the electron densitymaps, the boundary between the two protein molecules can be recognized, leading to thefollowing approximate molecular dimensions: 45×40×35A~3. Each molecule contains fourrod-shaped segments easily known as α-helices.     

The Crystal and Molecular Structure of Trichosanthin at 2.6 Resolution

The crystallographic refinement of trichosanthin has been performed at 2.6 resolution. The crystal and molecular structure of trichosanthin is described in detail in this paper. On summarizing the regularity of the amino acid sequences of eight kinds of ribosome inactivating proteins and combining with the crystal and molecular structure of trichosanthin, fifteen most conservative amino acid residues are analyzed. It is found that four most conservative polar amino acid residues Gln156, Glu160, Arg163 and Glu189 gather on the molecular surface on the boundary of the large and small domains, thus forming the active center of the protein molecule.     

THE COURSE OF THE POLYPEPTIDE CHAIN OF TRICHOSANTHIN MOLECULE

The course of the polypeptide chain of Trichosanthin has been determined from the electron density map at 4 resolution. The structure belongs to α+β type. It contains eight α-helices (39% of total residues) and four β-sheets (32% of total residues) which are made up of thirteen β-strands. The α-helices are relatively in the centre of the molecule and surrounded by β-sheets. This is the characteristic feature of Trichosanthin structure. This kind of structural arrangement has not been reported before. The transformation matrix and translation vector, which superpose two molecules in one asymmetric unit, were obtained. The root mean square error is 1.31 for this superposition.     

天花粉蛋白分子的肽链走向

本文证明从4分辨率的电子密度图上确定了一条合理的肽走向。天花粉蛋白属于α β型结构。整个分子包含八段α-螺旋,约占残基总数37％;13条β链组成4个β折叠层,占残基总数32％。α-螺旋相对地处于分子的中心,而4个β折叠层分布在外围。这是天花粉蛋白结构的显著特点,这种结构排布方式尚未见文献报道。用坐标叠合方法获得了不对称单位内两分子叠合时的旋转矩阵和平移向量,叠合后的均方根误差是1.31。