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Beckman L8-70M ultracentrifuge

Beckman L8-70M超速离心机

公司名称: Beckman Coulter
产品编号: L8-70M
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Expression and Purification of Yeast-derived GPCR, Gα and Gβγ Subunits for Structural and Dynamic Studies

In the last several years, as evidence of a surged number of GPCR-G complex structures, the expressions of GPCRs and G proteins for structural biology have achieved tremendous successes, mostly in insect and mammalian cell systems, resulting in more than 370 structures of over 70 GPCRs have been resolved. However, the challenge remains, particularly in the conformational transition and dynamics study area where a much higher quantity of the receptors and G proteins is required even in comparison to X-ray and cryo-EM (5 mg/ml, 3 μl/sample) when NMR spectroscopy (5 mg/ml, 250 μl /sample)

[摘要]  [摘要]在过去的几年中,作为GPCR-G复杂结构数量激增的证据,用于结构生物学的GPCR和G蛋白的表达已取得了巨大的成功,主要是在昆虫和哺乳动物细胞系统中,导致了370多个已解决了70多个GPCR的结构。但是,挑战仍然存在,特别是在构象转变和动力学研究领域,即使与X射线和冷冻EM相比(5 mg / ml,3μl /样品),也需要大量的受体和G蛋白。当应用NMR光谱法(5 mg / ml,250μl /样品)时。结果,i的表达水平 nsect和哺乳动物系统也很难满足这一需求,更不用说使用绝大多数系统使用这些系统生产GPCR和G蛋白的成本高昂了。因此,需要探索一种具有广泛适用性的有效,负担得起的实用方法。毕赤酵母表达系统已在GPCR制备中显示出其希望,并具有其他真核表达系统无法比拟的许多优点。在该系统中表达的GPCR价格便宜,易于操作,并且能够进行同位素标记。在此,我们提出最近开发并在我们的实验室升级的相关协议,包括表达和纯化的毕赤酵母衍生GPCR以G沿α和G βγ蛋白。我们预期这些协议将促进GPCR及其复合物的构象转变和动力学研究。

[背景] G蛋白偶联受体(GPCR)是最大的膜蛋白家族,在许多(病理)生理活动中起着关键作用。GPCR的或它们的效应物的功能障碍会导致各种病症,包括神经变性疾病,癌症,和慢性炎症(Overington等人,2006) ...

Isolation of Cytosol, Microsome, Free Polysomes (FPs) and Membrane-bound Polysomes (MBPs) from Arabidopsis Seedlings
[Abstract]  The plant endomembrane system plays vital roles for synthesis, modification and secretion of proteins and lipids. From the classic view, only mRNAs encoding secreted proteins could be targeted to the endoplasmic reticulum (ER) for translation via a co-translational translocation manner, however, recently this model has been challenged by accumulative evidence that lots of cytosolic mRNAs could also associate with ER, and that some categories of small RNAs are enriched on ER. These results suggested unrevealed functions of ER beyond our current knowledge. The large scale identification of RNAs ... [摘要]  植物内膜系统对蛋白质和脂质的合成,修饰和分泌起着至关重要的作用。 从经典观点来看,只有编码分泌蛋白质的mRNA才能通过协同翻译方式靶向内质网(ER)进行翻译,然而最近,这一模型已经被大量的细胞溶质mRNA也可能与 ER,并且一些类别的小RNA在ER上富集。 这些结果表明ER的功能超出了目前的知识。 在微粒体上大规模鉴定RNA和蛋白质对于显示ER功能至关重要,研究将由下一代测序技术提升。 该协议提供了从植物组织中分离细胞质,微粒体,游离多聚体(FP)和膜结合多聚体(MBP)的技术工作流程。 分离的级分适用于mRNA,小RNA和蛋白质的基因组广谱分析。
【背景】植物内膜系统对于细胞壁形成,脂质生物合成,蛋白质合成,修饰,折叠和贩运非常重要。根据共翻译易位模型,分泌蛋白N末端的信号肽由细胞溶质多核糖体合成,然后由ER上的信号识别粒子识别,其余蛋白质部分随后在ER上合成。根据该模型,只有编码分泌蛋白的mRNA可以被带到ER进行翻译(Peter和Johnson,1994)。然而,从哺乳动物和植物细胞ER(Lerner等人,2003; de ...

Preparation of Outer Membrane Vesicles from Myxococcus xanthus
[Abstract]  Outer membrane vesicles (OMVs) represent a unique sub-cellular compartment of bacteria that may act as a scaffold for various extracellular activities, including intercellular signaling. Myxococcus xanthus (M. xanthus) is a predatory bacterium that engages in cell-cell behaviors such as fruiting body formation and contact dependent lysis of other microbes. The OMVs of M. xanthus have been shown to have an elaborate architecture of chains and tubes that can connect cells within a biofilm. These higher order OMV structures have been shown to contain proteins exchanged ... [摘要]  外膜囊泡(OMV)代表细菌的独特亚细胞区室,其可以充当各种细胞外活性(包括细胞间信号传导)的支架。 (Myxococcus xanthus)是一种掠夺性细菌,其参与细胞 - 细胞行为,例如子实体形成和其他微生物的接触依赖性裂解。 M.M的OMV。 xanthus已经显示具有可以连接生物膜内的细胞的链和管的精细结构。 这些更高级的OMV结构已经显示含有交换为社区行为的蛋白质和具有抗生素活性的小分子,并且可以有助于促进定向交换。 M。 xanthus OMV允许相邻细胞之间的物质转移,用于运动和捕食。