• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Applied Microbiology >A Novel Glycosylphosphatidylinositol-Anchored Glycoside Hydrolase from Ustilago esculenta Functions in β-1,3-Glucan Degradation
【24h】

A Novel Glycosylphosphatidylinositol-Anchored Glycoside Hydrolase from Ustilago esculenta Functions in β-1,3-Glucan Degradation

机译：一种新的糖基磷脂酰肌醇锚定糖苷水解酶在β-1,3-葡聚糖降解中的作用

获取原文
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

A glycoside hydrolase responsible for laminarin degradation was partially purified to homogeneity from a Ustilago esculenta culture filtrate by weak-cation-exchange, strong-cation-exchange, and size-exclusion chromatography. Three proteins in enzymatically active fractions were digested with chymotrypsin followed by liquid chromatography-tandem mass spectrometry (LC/MS/MS) analysis, resulting in the identification of three peptide sequences that shared significant similarity to a putative β-1,3-glucanase, a member of glucoside hydrolase family 16 (GH16) from Sporisorium reilianum SRZ2. A gene encoding a laminarin-degrading enzyme from U. esculenta , lam16A , was isolated by PCR using degenerate primers designed based on the S. reilianum SRZ2 β-1,3-glucanase gene. Lam16A possesses a GH16 catalytic domain with an N-terminal signal peptide and a C-terminal glycosylphosphatidylinositol (GPI) anchor peptide. Recombinant Lam16A fused to an N-terminal FLAG peptide (Lam16A-FLAG) overexpressed in Aspergillus oryzae exhibited hydrolytic activity toward β-1,3-glucan specifically and was localized both in the extracellular and in the membrane fractions but not in the cell wall fraction. Lam16A without a GPI anchor signal peptide was secreted extracellularly and was not detected in the membrane fraction. Membrane-anchored Lam16A-FLAG was released completely by treatment with phosphatidylinositol-specific phospholipase C. These results suggest that Lam16A is anchored in the plasma membrane in order to modify β-1,3-glucan associated with the inner cell wall and that Lam16A is also used for the catabolism of β-1,3-glucan after its release in the extracellular medium.
机译：通过弱阳离子交换，强阳离子交换和尺寸排阻色谱法，将一部分负责层粘连蛋白降解的糖苷水解酶从Ustilago esculenta培养液中部分纯化至均质。用胰凝乳蛋白酶消化酶活性部分中的三种蛋白质，然后进行液相色谱-串联质谱（LC / MS / MS）分析，从而鉴定出与推定的β-1,3-葡聚糖酶具有显着相似性的三种肽序列， re孢孢霉SRZ2的糖苷水解酶家族16（GH16）的成员。使用基于re链霉菌SRZ2β-1,3-葡聚糖酶基因设计的简并引物，通过PCR分离出一种来自美国蓝藻的层板蛋白降解酶lam16A。 Lam16A具有GH16催化结构域，该结构域具有N端信号肽和C端糖基磷脂酰肌醇（GPI）锚定肽。融合到在米曲霉中过表达的N端FLAG肽（Lam16A-FLAG）融合的重组Lam16A表现出对β-1,3-葡聚糖的水解活性，并且既位于细胞外部分也位于膜部分中，但不在细胞壁部分中定位。没有GPI锚定信号肽的Lam16A在细胞外分泌，在膜级分中未检测到。通过用磷脂酰肌醇特异性磷脂酶C处理，膜锚定的Lam16A-FLAG完全释放。这些结果表明，Lam16A锚定在质膜上是为了修饰与内细胞壁相关的β-1,3-葡聚糖，而Lam16A是也用于β-1,3-葡聚糖在细胞外培养基中释放后的分解代谢。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号