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首页> 外文学位 >Production of chitosanase by recombinant Streptomyces lividans and enzymatic preparation of chitosan oligomers.
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Production of chitosanase by recombinant Streptomyces lividans and enzymatic preparation of chitosan oligomers.

机译：通过重组链霉菌链霉菌生产壳聚糖酶和酶促制备壳聚糖寡聚物。

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摘要

Elicitors are molecules which can trigger phytoalexin and pathogenesis-related proteins biosynthesis in plants. Chitosan as an elicitor has important roles in the interaction between pathogenic fungi and plants. Chitosan can inhibit fungal DNA transcription into mRNA, and it can also trigger the specific defensive genes which encode for at least 20 kinds of proteins related to resistant mechanism in plants.; Chitosanase is an endoglycosidase that can hydrolyze chitosan into oligosaccharide fragments. Chitosan heptamer-more fractions showed maximal activity in both antifungal properties and induction of plant resistance. In this work, recombinant strains of Streptomyces lividans were used for production of chitosanase, then we used this enzyme to prepare the active chitosan oligomers and to test their antifungal properties and their ability to induce defensive responses in plants.; In order to produce large amounts of chitosanase, the chitosanase gene from Streptomyces N174 was cloned in the high copy-number vector pFD666 and transformed into protoplasts of S. lividans TK24 and 10-164. These recombinant strains could produce chitosanase efficiently. The chitosanase activity could reach up to 95 units per millilitre of fermentation liquid on natural substrate (mycelium of Mucor rouxii. When DNA sequences were deleted in upstream or downstream of the chitosanase gene, chitosanase production became lower. Maybe, these sequences have some functions for the chitosanase gene expression and stability. In the chitosan medium, the chitosanase activity of the recombinant strains S. lividans TK24 decreased quickly after 3 days. We have shown that it related to the production of a specific proteolytic enzyme for chitosanase. On the contrary, the chitosanase activity could maintain high level after 3 days in M. rouxii mycelium as fermentation substrate. If S. lividans 10-164 was used as host for carrying the chitosanase gene, chitosanase activity reached high level after 2 days and the same level was maintained for a few days in D-glucosamine medium. The enzyme was recovered by polyacrylic acid precipitation. The enzyme prepared with this method has stable activity for long time.; Using chitosanase hydrolysis, the heptamer-more fractions were prepared. These chitosan oligomers could inhibit fungal growth and could induce the production of pathogenesis-related proteins such as {dollar}beta{dollar}-1,3-glucanase, chitinase and chitosanase by plants.
机译：激发子是可以触发植物中植物抗毒素和与病程相关的蛋白质生物合成的分子。壳聚糖作为引发剂在致病真菌和植物之间的相互作用中具有重要作用。壳聚糖可以抑制真菌DNA转录成mRNA，还可以触发特定的防御基因，该基因编码至少20种与植物抗性机制有关的蛋白质。壳聚糖酶是一种内切糖苷酶，可以将壳聚糖水解成寡糖片段。壳聚糖更多的七聚体级分在抗真菌特性和诱导植物抗性方面均显示出最大活性。在这项工作中，使用链霉菌链霉菌的重组菌株生产壳聚糖酶，然后我们使用该酶制备活性壳聚糖寡聚物，并测试其抗真菌特性以及诱导植物防御反应的能力。为了产生大量的壳聚糖酶，将来自链霉菌N174的壳聚糖酶基因克隆到高拷贝数载体pFD666中，并转化为S.lividans TK24和10-164的原生质体。这些重组菌株可以有效地产生壳聚糖酶。在自然底物（Mucor rouxii的菌丝体）上，每毫升发酵液中的壳聚糖酶活性最高可达到95个单位。当在壳聚糖酶基因的上游或下游缺失DNA序列时，壳聚糖酶的产量会降低，也许这些序列具有一定的功能。在壳聚糖培养基中，重组菌株S. lividans TK24的壳聚糖酶活性在3天后迅速下降，这表明它与壳聚糖酶的特定蛋白水解酶的产生有关。肉毒杆菌菌丝体作为发酵底物的壳聚糖酶活性在3天后仍能保持较高水平，如果以链霉菌10-164为宿主携带壳聚糖酶基因，则在2天后壳聚糖酶活性达到较高水平，并保持在同一水平。在D-葡萄糖胺培养基中放置数天，然后通过聚丙烯酸沉淀回收该酶，用该方法制得的酶od具有长期稳定的活性。使用壳聚糖酶水解，制备了更多的七聚体级分。这些壳聚糖低聚物可以抑制真菌生长，并可以诱导植物与病程相关的蛋白质的产生，例如{beta} {{dollar} -1,3-葡聚糖酶，几丁质酶和几丁质酶。

著录项

  • 作者

    Li, Tong.;

  • 作者单位

    Universite de Sherbrooke (Canada).;

  • 授予单位 Universite de Sherbrooke (Canada).;
  • 学科 Biology Microbiology.
  • 学位 M.Sc.
  • 年度 1993
  • 页码 117 p.
  • 总页数 117
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 微生物学;
  • 关键词

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