X射线衍射晶体法解析脱卤酶DehDIV-R结构的研究

R-2-卤代酸脱卤酶能立体选择性水解R-2-卤代酸。解析酶的单晶结构对提高酶的选择性和活性提供了直接的结构指导,是目前酶结构领域研究的前沿。以实验室前期得到的来自假单胞菌ZJU26的R-2-氯丙酸脱卤酶(Deh DIV-R)为研究对象,采用X射线衍射晶体法进行结构解析。采用pp SUMO载体融合表达Deh DIV-R蛋白,依次通过Ni-NTA亲和层析、透析酶切、二次NiNTA亲和层析以及凝胶过滤层析纯化得到单一条带,且均一性好的蛋白。接着对结晶条件进行初筛与优化,得到的最佳结晶条件为0.1mol/L HEPES p H 7,12%PEG 6 000,0.2mol/L Mg Cl2,8mmol/L CHAPS。晶体在上海同步辐射光源BL18U1线站上收集衍射数据,采用分子置换法成功解析获得了分辨率为2.35的Deh DIV-R的晶体结构。Ramachandran图表明98.02%的氨基酸位于最适区,证明了该结构的合理性。Deh DIV-R的纯化、结晶以及结构解析为进一步深入了解其结构和功能奠定了基础。     

灰雁(Anser anser)氧合血红蛋白的晶体生长和X射线衍射的初步分析

灰雁血红蛋白分子与斑头雁血红蛋白分子结构相近,二者对比有助于分析斑头雁血红蛋白的高氧亲和性机理。本文报道了以聚乙二醇６,０００为沉淀剂生长出灰雁氧合血红蛋白的晶体,经Ｘ射线衍射分析确定,晶体属单斜晶系,空间群为Ｐ２１,晶胞参数为ａ＝５．７５５２ｎｍ,ｂ＝８．０６２４ｎｍ,ｃ＝７．２５７１ｎｍ,α＝９０°,β＝１０２．７５°,γ＝９０°,晶胞体积为３．２８×１０２ｎｍ３,每个不对称单位中包含一个四聚体分子。研究灰雁氧合血红蛋白的晶体结构,可以得到较准确的结构信息。     

X-射线纤维衍射在多糖构型分析中应用的研究进展

本文从Ｘ射线纤维衍射法的原理,实验方法及应用实例三个方面对该方法在多糖构型分析中的应用状况进行综述,引用文献２０篇。     

Co2+-Tyr配合物的合成及X射线衍射分析

研究了Co2+-Tyr配合物的合成方法,探讨了合成工艺的主要影响因素,确定了最佳反应时间、温度、pH值及原料配比,采用X射线衍射光谱及元素分析对配合物结构进行了分析鉴定,结果表明Co2+能与Tyr形成配合物。     

斑头雁(Anser indicus)高铁血红蛋白的晶体生长和初步的X-射线衍射研究

本文报导了斑头雁(Auser indicus)高铁血红蛋白的晶体生长和初步的晶体学研究.晶体的衍射分辨率为2.5A或更好,四方晶系.空间群为P4:2_12,晶胞参数为:a=b=80.8A,e=107.3A,V=700523A~3.每个晶胞中含4个四聚体分子,每个不对称单位中含半个四聚体分子.     

综述: X射线晶体衍射是解析出青蒿素三维结构的唯一方法

X射线晶体衍射是解析出青蒿素三维结构的唯一方法     

对X射线在医学影像中的影响研究

人们利用X线穿透能力强的特点,根据人体不同的组织对X线的吸收程度不同,均匀的X线穿透人体组织后,其不均匀的分布其实就是人体组织的投影。把这种成像技术应用在医学上,就可以得到患病处的位置信息。文章介绍了X线的原理及在医学影像上的作用。     

北京鸭(Anas platyrhynchos)氧合血红蛋白的晶体生长和X射线晶体学的初步研究

北京鸭氧合血红蛋白晶体在以聚乙二醇６０００为沉淀剂,在ｐＨ７．１,血红蛋白浓度为２５ｍｇ／ｍｌ条件下采用一批结晶法培养得到,并且适合于Ｘ射线衍射分析。经Ｘ射线平面探测仪收集衍射强度数据和处理,确定晶体属四方晶系,空间群为Ｐ４２２１２,晶胞参数为ａ＝ｂ＝８．１１３ｎｍ,ｃ＝１０．７０７ｎｍ,α＝β＝γ＝９０°,每个不对称单位中包含半个四聚体分子。进一步得到北京鸭氧合血红蛋白高分辨率晶体结构,通过与其高海拔同类－斑头雁分析比较深入研究斑头雁血红蛋白分子的高氧亲和性机制。     

粉末X射线衍射图谱计算植物淀粉结晶度方法的探讨

植物淀粉有A-型、B-型和C-型3种晶体。以水稻(Oryza sativa)、马铃薯(Solanum tuberosum)、豌豆(Pisum sativum) 和莲藕(Nelumbo nucifera)淀粉为材料, 利用粉末X-射线衍射仪(XRD)调查了不同晶体类型淀粉的波谱特征, 探讨XRD波谱相对结晶度的计算方法。软件峰拟合法、软件曲线法、直线作图法和曲线作图法均可用于计算淀粉XRD波谱的相对结晶度, 以曲线作图法计算结果较为可靠。利用曲线作图法得出的结果表明, 稻米淀粉的结晶度与直链淀粉含量呈显著线性负相关, 酸解莲藕淀粉的结晶度与淀粉酸水解度呈显著线性正相关。酸水解使莲藕淀粉的C-型晶体转变为A-型晶体。上述研究结果为利用XRD分析植物淀粉晶体类型和计算相对结晶度提供了重要参考。     

孕烷X受体配体结合结构域蛋白晶体的X线衍射结构解析

目的:利用X线衍射技术解析孕烷X受体(PXR)配体结合结构域(LBD)蛋白晶体的3维结构。方法:对PXR蛋白LBD(130~434氨基酸残基)序列进行密码子优化并化学合成后克隆至pRSFDuet-1表达载体,再将载体导入大肠杆菌BL21(DE3),对PXR-LBD蛋白进行原核表达与分离纯化;采用晶体筛选试剂盒筛选蛋白结晶条件,采用悬滴法获得目标蛋白的晶体;对获得的蛋白晶体进行X线晶体衍射检测,并收集相关数据建立PXR-LBD的三维结构。结果:获得了PXR-LBD的高质量晶体并利用X线衍射解析了该蛋白质晶体的结构数据,使用Phenix.refine软件和COOT软件等对结构进行修正,最终获得了高分辨率的3维结构数据。结论:完成了孕烷X受体配体结合结构域蛋白晶体的X线衍射结构解析,为研究和开发PXR相关药物奠定了基础。     

X射线衍射法表征微量元素钴-半胱氨酸配合物

研究了钴-半胱氨酸配合物的合成方法,通过大量的实验探讨了温度、原料配比、反应时间以及pH对反应的影响,从而确定了最佳的合成条件。采用X射线衍射光谱及元素分析对配合物结构进行了分析鉴定,结果表明钴能与半胱氨酸形成配合物。     

X-Ray Diffraction of Iron Containing Samples: The Importance of a Suitable Configuration

In X-ray diffraction, a good combination of configuration and sample is essential. Copper radiation for iron containing materials leads to a high background. Although this has been recognized, many researchers still use this combination. To clearly show the unsuitability of copper radiation for iron oxides, magnetite, goethite, maghemite, and hematite were analysed in different configurations using copper or cobalt radiation. Results show effects of fluorescence repressing measures and different radiation sources. Copper radiation diffractograms make phase identification contestable. Studies using copper radiation for iron oxides must therefore be carefully evaluated. Cobalt radiation yielded high quality diffractograms, making phase identification unambiguous.     

Crystallization and Preliminary X-Ray Diffraction Analysis of the 190-Å-Long Coiled-Coil Dimerization Domain of the Actin-Bundling Protein Cortexillin I fromDictyostelium discoideum

We have crystallized the ≈190-Å-long parallel two-stranded coiled-coil oligomerization domain of the actin-bundling protein cortexillin I fromDictyostelium discoideum. The orthorhombic crystals belong to the space group C222₁with unit cell dimensions ofa= 71.3 Å,b= 127.8 Å, andc= 91.6 Å. As both native and selenomethionine-substituted protein crystals diffract to 3.0 and 2.85 Å resolution, respectively, using synchrotron radiation, they are suitable for the first high-resolution structural analysis of a two-stranded coiled coil comprising more than six heptad repeats. Moreover, because the polypeptide chain fragment contains a recently identified two-heptad-repeat long sequence that is indispensable for the assembly of the cortexillin I coiled-coil oligomerization domain, its high-resolution structure should enable us to extend our knowledge on the molecular mechanisms underlaying coiled-coil formation and to establish the precise manner in which the two “trigger” sequences interact with one another in the dimer.     

Dynamic Light Scattering Application to Study Protein Interactions in Electrolyte Solutions

The concentration dependence of the diffusion coefficient of particles suspended in solution depends primarily on the occupied volume fraction and on repulsive and attractive forces. This dependency is expressed by the interaction parameter, which can be assessed experimentally by light scattering measurements and have been determined for the diffusion coefficient of BSA under different salt concentration conditions in the present work. The result shows that the diffusion coefficient of protein grows up with increasing protein concentration, and when the ionic strength turns up gradually the diffusion coefficient decreases with protein concentrations increasing. The concentration dependence of BSA diffusion coefficients is interpreted in the context of a two-body potential of mean force, which includes repulsive hard-sphere and Coulombic interactions and attractive dispersion. With the increase of ionic strength, Debye screening decreases, protein interaction changes from repulsion to attraction, and protein begins to aggregate. By means of the concentration dependence of BSA diffusion coefficients, one can obtain the parameters of protein interactions and can find that protein bears a net effective charge of –9.0 e and has a Hamaker constant of 2.8k_BT. This work demonstrates that DLS is an effective technique of studying protein interactions.     

A Rare Lysozyme Crystal Form Solved Using Highly Redundant Multiple Electron Diffraction Datasets from Micron-Sized Crystals

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Crystallization and preliminary X-ray crystallographic analysis of ImmE7 protein of colicin E7

The ImmE7 protein, which can bind specifically to the DNase colicin E7 and neutralize its bactericidal activity, has been purified and crystallized in two different crystal forms by vapor diffusion method. The orthorhombic crystals belong to space group I222 or I2₁2₁2₁ and have unit cell dimensions a = 75.1 Å, b = 50.5 Å, and c = 45.4 Å. The second form is monoclinic space group P2₁ with ceil dimensions a = 29.3 Å, b = 102.7 Å, c = 53.0 Å and β = 91.5°. The orthorhombic crystals diffract to 1.8 Å resolution, and are suitable for high-resolution X-ray analysis. © 1995 Wiley-Liss, Inc.     

Quantitative Locomotion Study of Freely Swimming Micro-organisms Using Laser Diffraction

Soil and aquatic microscopic organisms live and behave in a complex three-dimensional environment. Most studies of microscopic organism behavior, in contrast, have been conducted using microscope-based approaches, which limit the movement and behavior to a narrow, nearly two-dimensional focal field.¹ We present a novel analytical approach that provides real-time analysis of freely swimming C. elegans in a cuvette without dependence on microscope-based equipment. This approach consists of tracking the temporal periodicity of diffraction patterns generated by directing laser light through the cuvette. We measure oscillation frequencies for freely swimming nematodes.Analysis of the far-field diffraction patterns reveals clues about the waveforms of the nematodes. Diffraction is the process of light bending around an object. In this case light is diffracted by the organisms. The light waves interfere and can form a diffraction pattern. A far-field, or Fraunhofer, diffraction pattern is formed if the screen-to-object distance is much larger than the diffracting object. In this case, the diffraction pattern can be calculated (modeled) using a Fourier transform.²C. elegans are free-living soil-dwelling nematodes that navigate in three dimensions. They move both on a solid matrix like soil or agar in a sinusoidal locomotory pattern called crawling and in liquid in a different pattern called swimming.³ The roles played by sensory information provided by mechanosensory, chemosensory, and thermosensory cells that govern plastic changes in locomotory patterns and switches in patterns are only beginning to be elucidated.⁴ We describe an optical approach to measuring nematode locomotion in three dimensions that does not require a microscope and will enable us to begin to explore the complexities of nematode locomotion under different conditions.