Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/23788
標題: Paenibacillus sp. 葡萄聚醣水解酵素之純化與功能分析
Purification and Function Analysis of beta-1,3-Glucanase from Paenibacillus sp.
作者: 陳俊宇
Chen, Jiun-Yu
關鍵字: Paenibacillus sp.;Paenibacillus sp.;beta-1;3-Glucanase;葡萄聚醣水解酵素
出版社: 生物化學研究所
摘要: 
中文摘要
在自然界中有些真菌會對植物會造成病害,造成對農作物相當大的損失,為了防治病害,若使用大量的化學藥劑,會對環境造成污染,所以我們希望研究其它防治的方法來取代化學農藥。本試驗嘗試以生物防治的觀念,利用具高活性β-1,3-glucanase的微生物與環境中的病原真菌拮抗,達到防治作物之真菌病害,或降低作物栽培時化學藥劑之使用量。
本試驗篩選得到Paenibacillus sp.具有分泌β-1,3-glucanase的能力,在試驗室經由LB培養基加入pachyman誘導培養,經由Q Sepharose 及SP Sepharose純化,得到β-1,3-glucanase的分子量約30 kDa,酵素反應最適當反應的pH 為5.0,在以laminarin (可溶性 b-1,3-glucan)當作受質在pH 5.0溫度為50℃有最高活性,此酵素對加熱處理的lichenan (b-1,3-1,4-glucan)有比laminarin更高的活性,酵素對不溶性b-1,3-glucan的水解能力很低,對其它不溶性受質則沒有活性,但是酵素對一些不溶性受質有不錯的結合能力,酵素可能具有cellulose-binding domains,對cellulose與xylan有很好的結合能力。對真菌的抑制實驗上,將病原性真菌Rhizoctonia solani AG-4與β-1,3-glucanase (150 mg/ml, 10 mM [pH 6.0]磷酸緩衝溶液)於室內培養,發現對病原性真菌Rhizoctonia solani AG-4的生長有明顯抑制的效果,並造成原生質外流及菌絲的萎縮的現象。

Abstract
A beta-1,3-glucanase was isolated from the culture supernatant of a Paenibacillus sp. grown on L broth medium containing 1 g pachyman (an insoluble b-1,3-glucan). The molecular mass of the enzyme was estimated to be 30-kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The 30-kDa emzyme was purified to homogeneity from the culture supernatant in this study. The property of this enzyme was examined. The enzyme belongs to a category of endo type 1,3-glucan glucanohydrolases. This enzyme was active at pH 5.0. The enzyme had activity on some insoluble beta-1,3-glucan such as curdlan, pachyman, Zymaose A, lichenan and no activity on others insoluble polysaccharide, but this enzyme bond to curdlan, cellulose and xylan. The optimum temperature of the reactions was 50℃when laminarin (a soluble beta-1,3-glucan) was used as the substrate at pH 5.0. The enzyme hydrolyzed lichenan (beta-1,3-1,4-glucan) more effectively than laminarin. A b-1,3-glucan, the substrate for the enzyme is a major component of the cell walls of pathogenic and potentially pathogenic fungi. The enzyme was lysed effecively fungal (Rhizoctonia solani AG-4) cell walls and resulted in the delay of fungi growth.
URI: http://hdl.handle.net/11455/23788
Appears in Collections:生物化學研究所

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