摘要
从米曲霉（Aspergillus oryzae FAFU）发酵产物中提取淀粉酶,通过硫酸铵盐析、透析、DEAE-Sepharose离子交换层析和Sephacryl S-200凝胶柱层析等纯化步骤,回收活性组分,结合十二烷基磺酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）进行纯度鉴定,共分离纯化出3种酶：酶Ⅰ、酶Ⅱ、酶Ⅲ。通过酶解淀粉实验,酶解产物经薄层层析（TLC）分析,鉴定出酶Ⅰ、酶Ⅱ为液化酶,酶Ⅲ为糖化酶,经SDS-PAGE电泳确定分子量大小分别为：48.6、49.7、93.5 k Da。对酶Ⅰ、酶Ⅱ的酶学性质进行初步的探究：酶Ⅰ的最适反应温度为50℃,温度在50℃以下稳定性好,55℃的半衰期是40 min,最适反应p H为6.0,在p H7.2~9.6范围内稳定性好;酶Ⅱ的最适反应温度为50℃,温度在45℃以下稳定性好,50℃的半衰期是30 min,最适反应p H为6.6,在p H8.6~10.0范围内稳定性好。
The crude amylase was extracted from Aspergillus oryzae FAFU. The enzymes were purified by precipi- tation through ammonium sulfate, desahed by dialysis, and successively purified by chromatography on DEAE Sepha- rose and Sephacryl S-200 column to produce three amylase fractions termed Amy-I,Amy-II, and Amy-Ⅲ, respective- ly. Amy- I and Amy- Ⅱ were identified as α-amylase, Amy-Ⅲ was identified as glucoamylase according to the results of TLC analysis for their enzymatic hydrolysates, and their molecular mass were about 48.6, 49.7, 93.5 kDa respec- tively through SDS-PAGE. Some enzymatic properties of Amy-I and Amy-II were characterized in this research. The optimal reaction temperature of Ant-I was determined to be 50 ℃. It was stable when the temperature was lower than 50 ℃ , and its half-life was 40 min at 55 ℃. The optimal reaction pH was 6.0, and its pH stabilities were kept in the range 7.2 - 9.6. The optimal reaction temperature of Amy- 11 was determined to be 50℃, it was stable when the tem- perature lower than 45℃ and its half-life was 30 min at 50℃. Its optimal reaction pH was 6.6, and it kept stable in the pH range 8.6 - 10.0.
出处
《食品与发酵工业》
CAS
CSCD
北大核心
2016年第11期71-76,共6页
Food and Fermentation Industries
基金
中国博士后科学基金第九批特等资助项目(2016T90591)
中国博士后科学基金第57批面上一等资助项目(2015M570549)
福建省自然科学基金计划面上项目(2016J01095)
关键词
米曲霉
淀粉酶
分离纯化
酶学特性
Aspergillus oryzae FAFU
amylase
purification
characteristics
作者简介
硕士研究生
（吕旭聪博士为通讯作者，E-mail：xu-cong1154@qq．com）
