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50 µL, Model 705 N SYR, Cemented NDL, 22s ga, 2 in, point style 2

Company: Hamilton
Catalog#: 80500
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Xenopus laevis Oocytes Preparation for in-Cell EPR Spectroscopy
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Date:
2018-04-05
[Abstract]  One of the most exciting perspectives for studying bio-macromolecules comes from the emerging field of in-cell spectroscopy, which enables to determine the structure and dynamics of bio-macromolecules in the cell. In-cell electron paramagnetic resonance (EPR) spectroscopy in combination with micro-injection of bio-macromolecules into Xenopus laevis oocytes is ideally suited for this purpose. Xenopus laevis oocytes are a commonly used eukaryotic cell model in different fields of biology, such as cell- and development-biology. For in-cell EPR, the bio-macromolecules of interest are microinjected into the Xenopus laevis oocytes upon site-directed spin labeling. The sample solution is filled into a thin glass capillary by means of Nanoliter Injector and after that ... [摘要]  研究生物大分子的最令人兴奋的观点之一来自于新兴的细胞内光谱学领域,它能够确定细胞中生物大分子的结构和动力学。细胞内电子顺磁共振(EPR)光谱结合将生物大分子微注射到非洲爪蟾卵母细胞中非常适合于此目的。非洲爪蟾卵母细胞是生物学不同领域常用的真核细胞模型,如细胞和发育生物学。对于细胞内EPR,感兴趣的生物大分子通过定点自旋标记显微注射到非洲爪蟾卵母细胞中。通过Nanoliter注射器将样品溶液填充到薄玻璃毛细管中,然后通过小心地穿刺薄膜将其微注射入非洲爪蟾卵母细胞的黑色动物部分。之后,取决于最终的细胞内EPR实验的种类,将三个或五个显微注射的非洲爪蟾卵母细胞装载到Q波段EPR样品管中,随后进行任选的休克冷冻(用于实验冷冻溶液)并且在期望的温育时间之后测量(在低温或生理温度下)。由于显微注射样品的细胞毒性作用和顺磁性自旋标记在还原性细胞环境中的稳定性,孵育时间受到限制。通过监测细胞形态和减少动力学来量化这两个方面。

【背景】电子顺磁共振(EPR)光谱学是用于表征顺磁系统的选择方法(Atherton,1993; Gerson等人,1994; Jeschke和Schweiger,2001)。反磁性生物大分子可以通过定点自旋标记(SDSL)进行EPR光谱学分析,通常使用氮氧化物作为自旋标记(Hubbell和Altenbach,1994; Feix和Klug,2002; ...

Assay to Measure Interactions between Purified Drp1 and Synthetic Liposomes
Author:
Date:
2017-05-05
[Abstract]  A mitochondrion is a dynamic intracellular organelle that actively divides and fuses to control its size, number and shape in cells. A regulated balance between mitochondrial division and fusion is fundamental to the function, distribution and turnover of mitochondria (Roy et al., 2015). Mitochondrial division is mediated by dynamin-related protein 1 (Drp1), a mechano-chemical GTPase that constricts mitochondrial membranes (Tamura et al., 2011). Mitochondrial membrane lipids such as phosphatidic acid and cardiolipin bind Drp1, and Drp1-phospholipid interactions provide key regulatory mechanisms for mitochondrial division (Montessuit et al., 2010; Bustillo-Zabalbeitia et al., 2014; Macdonald et al., 2014; Stepanyants et al., 2015; ... [摘要]  线粒体是一种动态的细胞内细胞器,主动分裂和融合以控制细胞的大小,数量和形状。线粒体分裂和融合之间的调节平衡是线粒体功能,分布和周转的基础(Roy等,2015)。线粒体分化是由动力蛋白相关蛋白1(Drp1)介导的,其是限制线粒体膜的机械化学GTP酶(Tamura等人,2011)。线粒体膜脂质如磷脂酸和心磷脂结合Drp1,并且Drp1磷脂相互作用提供线粒体分裂的关键调控机制(Montessuit等人,2010; Bustillo-Zabalbeitia等人2014年; Macdonald等人,2014年; Stepanyants等人,2015; Adachi等人,2016)。在这里,我们描述了使用纯化的重组Drp1和具有定义的一组磷脂的合成脂质体定量测量Drp1与脂质的相互作用的生物化学实验。该测定使得可以定义蛋白质 - 脂质相互作用的特异性以及头基和酰基链的作用。

背景 蛋白质和膜脂质的相互作用对于细胞如细胞器分裂中生物膜的重塑至关重要。在线粒体分裂中,Drp1限制线粒体膜并驱动该膜重塑过程。我们最近显示,信号磷脂,磷脂酸与Drp1相互作用,并通过限制线粒体上的组装分裂机制(Adachi等人,2016)产生启动步骤。 Drp1识别磷脂酸的头基和酰基链。为了分析Drp1-磷脂酸结合,我们建立了几种蛋白质 - ...

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