| Extraction of Intracellular and Cell Wall Proteins from Leaves and Roots of Harsh Hakea
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Author:
Date:
2015-12-05
[Abstract] Plant proteins can be targeted to intracellular (i.e., cytosol, vacuole, organelles etc.) or extracellular (i.e., cell walls, apoplast) compartments. Dual targeting is a key mechanism with important implications for plant metabolism, growth, development and defense etc. Harsh Hakea (Hakea prostrata R.Br.) is a perennial species and member of the Proteaceae family that thrives on extremely phosphate impoverished soils of southwestern Australia. Harsh Hakea is not a common model organism, but has been widely developed for physiological and molecular/biochemical studies of the endogenous adaptations of an ‘extremophile’ plant species to abiotic stress, including low phosphorus tolerance. Tissues of Harsh Hakea contain large amounts of compounds (e.g. ...
[摘要] 植物蛋白可以靶向细胞内（即胞质溶胶，液泡，细胞器等）或细胞外（即细胞壁，质外体）区室。双重靶向是对植物新陈代谢，生长，发育和防御等具有重要影响的关键机制。 Harsh Hakea（ Hakea prostrata R.Br.）是一种多年生物种和成员的泛革兰科家族，在澳大利亚西南部的极端磷酸盐贫困土壤上生长。 Harsh Hakea不是一种常见的模式生物，而是广泛开发用于"极端"植物物种对非生物胁迫（包括低磷耐性）的内源性适应的生理学和分子/生物化学研究。 Harsh Hakea的组织含有大量干扰可溶性蛋白质提取的化合物（例如，酚类化合物）。我们以前优化了来自Harsh Hakea蛋白质组织根的细胞内蛋白质的提取，将可溶性蛋白质产量提高至少10倍（Shane等人，2013）。在这里，我们描述的提取和细胞内分离从"松散绑定"细胞壁蛋白质在Harsh Hakea的协议。
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| Protein Sample Preparation for Proteomic Analysis in Leishmania donovani
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Author:
Date:
2014-03-05
[Abstract] Leishmania is a genus of trypanosomatid protozoa and is the parasite responsible for the disease leishmaniasis. These protozoa, regulate their gene expression in an atypical way, compared to other higher eukaryotes. The regulation of gene expression is characterized by a predominance of post-transcriptional over pre-transcriptional regulatory mechanisms (Clayton, 2002). Thus proteomic analysis has proven an essential tool for understanding pathways implicated in Leishmania infectivity, host-parasite interactions, drug resistance and others. When employing a comparative proteomics analysis between different parasitic cell lines, it is essential that these lines are cultivated in exactly the same way, in the same cell density and growth phase. More importantly when ...
[摘要] 利什曼原虫属是锥虫病原生动物属，是负责疾病利什曼病的寄生虫。这些原生动物，与其他高等真核生物相比，以非典型方式调节它们的基因表达。基因表达的调节的特征在于转录后优先于转录前调节机制（Clayton，2002）。因此，蛋白质组学分析已经证明是了解利什曼原虫感染，宿主寄生虫相互作用，耐药性和其他相关途径的重要工具。当在不同寄生细胞系之间使用比较蛋白质组学分析时，以相同的方式，在相同的细胞密度和生长期中培养这些细胞系是必要的。更重要的是，当怀疑细胞周期缺陷时，必须在同一细胞周期阶段同步细胞系，以消除可能的人工因素。该方案描述了在杜氏利什曼原虫（ donovani ）中用于蛋白质组分析的全蛋白样品的制备。
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| Protein Extraction, Acid Phosphatase Activity Assays, and Determination of Soluble Protein Concentration
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Author:
Date:
2013-09-05
[Abstract] Acid phosphatases (APases) catalyze the hydrolysis of inorganic phosphate (Pi) from a broad range of Pi-monoesters with an acidic pH optimum. The liberated Pi is reassimilated into cellular metabolism via mitochondrial or chloroplastic ATP synthases of respiration or photosynthesis, respectively. Eukaryotic APases exist as a wide variety of tissue- and/or cellular compartment-specific isozymes that display marked differences in their physical and kinetic properties. Increases in intracellular (vacuolar) and secreted APase activities are useful biochemical markers of plant nutritional Pi deficiency. The protocols for protein extraction, APase activity determination and measurement of soluble protein concentration from plant tissues or cell suspension cultures are presented.
[摘要] 酸性磷酸酶（APase）催化来自具有酸性pH最佳值的宽范围的P 1 - 单酯的无机磷酸盐（P 1）的水解。 释放的Pi分别通过呼吸或光合作用的线粒体或叶绿体ATP合酶再吸收到细胞代谢中。 真核APase作为多种组织和/或细胞区室特异性同工酶存在，其在它们的物理和动力学性质上显示出显着的差异。 细胞内（液泡）和分泌的APase活性的增加是植物营养缺乏的有用的生物化学标记物。 提出了蛋白质提取，APase活性测定和来自植物组织或细胞悬浮培养物的可溶性蛋白质浓度的测定方案。
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