| Quantification of Chitinase Activity in Fusarium oxysporum
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Author:
Date:
2016-08-05
[Abstract] Fungal morphogenetic development requires modification and plasticity of the cell wall, which implies synthesis and remodelling of its components, including chitin and glucan. Thus chitinase and glucanase activities are crucial for cell-wall biogenesis and cell division. Quantification of chitinase activity might be useful to identify structural defects that could negatively influence growth and morphogenesis of some filamentous fungi like Fusarium oxysporum, which produces both intracellular and secreted chitinases. The chitinolytic enzymes are categorized based on their enzymatic action on chitin substrates. Endochitinases are defined as the enzymes catalyzing the random cleavage at internal points in the chitin chain. Exochitinases catalyze the progressive release of ...
[摘要] 真菌形态发生需要细胞壁的修饰和可塑性，这意味着其组分（包括壳多糖和葡聚糖）的合成和重塑。因此几丁质酶和葡聚糖酶活性对于细胞壁生物发生和细胞分裂是至关重要的。几丁质酶活性的定量可能有助于鉴定可能负面影响一些丝状真菌如产生细胞内和分泌型几丁质酶的尖孢镰刀菌的生长和形态发生的结构缺陷。基于它们对壳多糖底物的酶作用将壳聚糖分解酶分类。内酯酶被定义为催化几丁质链中内部点的随机裂解的酶。外切球蛋白酶催化乙酰壳寡糖或N-乙酰葡糖胺从几丁质的非还原端逐步释放，因此分别称为壳聚糖酶和β-N-乙酰氨基葡糖苷酶。在这里，我们描述了一种简单的方法来轻易地纯化几丁质酶，以便比较不同的F的内切几丁质酶活性和外切几丁质酶活性。尖孢镰孢菌株。该方案可适用于任何真菌物种。
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| Determination of Intra- and Extracellular Glucose in Mycelium of Fusarium oxysporum
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Author:
Date:
2016-07-20
[Abstract] To study alterations in the metabolism and/or in the transport of glucose during Fusarium oxysporum vegetative growth, we determined intracellular glucose levels in different fungal strains, as well as the amount of glucose remaining in the supernatants after growth in synthetic medium (SM) supplemented with either 0.05 or 2.5% glucose. We used the Glucose (GO) Assay Kit (Sigma-Aldrich) following the instructions of the manufacturer with some modifications. The protocol described here can be applied to other filamentous fungi.
[摘要] 为了研究尖镰孢营养生长期间葡萄糖的代谢和/或转运中的改变，我们测定了不同真菌菌株中的细胞内葡萄糖水平以及生长后上清液中剩余的葡萄糖量 在补充有0.05或2.5％葡萄糖的合成培养基（SM）中。 我们使用葡萄糖（GO）测定试剂盒（Sigma-Aldrich）按照制造商的指示进行一些修改。 这里描述的协议可以应用于其他丝状真菌。
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| Shikimate Hydroxycinnamoyl Transferase (HCT) Activity Assays in Populus nigra
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Author:
Date:
2013-11-20
[Abstract] Lignin is a complex phenolic polymer deposited in secondarily-thickened plant cell walls. The polymer is mainly derived from the three primary monolignols: p-coumaryl, coniferyl and sinapyl alcohol which give rise to p-hydroxyphenyl, guaiacyl and syringyl units (H, G and S units, respectively) when coupled into the polymer. The building blocks differ in their degree of methoxylation and their biosynthetic pathway is catalyzed by more than 10 enzymes. HCT plays a crucial role by channeling the phenylpropanoids towards the production of coniferyl and sinapyl alcohols. Interestingly, HCT has been reported to be implicated in the pathway both upstream and downstream of the 3-hydroxylation of the aromatic ring of p-coumaroyl shikimate (Figure 1) (Hoffmann et al., ...
[摘要] 木质素是沉积在二次增厚的植物细胞壁中的复杂酚类聚合物。聚合物主要衍生自当偶联时产生对羟基苯基，愈创木基和紫丁香基单元（分别为H，G和S单元）的三种主要单独的木质素：p - 香豆酰基，松柏基和芥子醇进入聚合物。结构单元的甲氧基化程度不同，它们的生物合成途径被超过10种酶催化。 HCT通过将苯丙素类导向松柏醇和芥子醇的生产来发挥关键作用。有趣的是，据报道，HCT涉及p - 香豆酰基莽草酸的芳环的3-羟基化的上游和下游途径（图1）（Hoffmann等， 2003; Hoffmann等人，2004; Vanholme等人，2013b）。这些特征突出了开发在植物中可靠地测量HCT活性的测定法的重要性。在这里，我们描述了基于UPLC-MS的方法，用于分析黑曲霉的木质部总蛋白提取物中的HCT活性，其可以适用于其他木本和草本植物物种。该方案最初在Vanholme等人（2013a）中描述。
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