α-淀粉酶产生菌的筛选鉴定及Ba8a蛋白的酶学特性

余振; 徐玉荣; 吴哲宽; 张静怡; 张小宇; 俸婷; 程国军; 李晓华; 饶雄飞

doi:10.13300/j.cnki.hnlkxb.2025.04.014

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α-淀粉酶产生菌的筛选鉴定及Ba8a蛋白的酶学特性

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- α-淀粉酶产生菌的筛选鉴定及Ba8a蛋白的酶学特性
- Isolation and identification of α-amylase producing bacteria and enzymatic characteristics of Ba8a protein
- 华中农业大学学报 2025年44卷第4期页码：143-149
- 作者机构：
  
  1.湖北省烟草科学研究院，武汉 430030
  2.中南民族大学生命科学学院/ 微生物资源与利用湖北省工程技术研究中心，武汉 430074
- 作者简介：
  
  余振，E-mail：304226690@qq.com
  李晓华，E-mail：lixiaohua@mail.scuec.edu.cn
  饶雄飞，E-mail：raoxiongfei@163.com
- 基金信息：
  
  中国烟草总公司“雪茄烟叶开发与应用重大专项”揭榜棒挂帅项目（中烟办2022［111］号,110202201039（XJ-10））;国家自然科学基金项目(31070087;32372809);湖北省大学生创新创业训练计划项目(SCX2024066);余振,E-mail：304226690@qq.com
- DOI：10.13300/j.cnki.hnlkxb.2025.04.014
  中图分类号： S476.1
- 收稿：2024-09-25，
  
  纸质出版：2025-07-05
- 稿件说明：
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余振，徐玉荣，吴哲宽，等.α-淀粉酶产生菌的筛选鉴定及Ba8a蛋白的酶学特性［J］.华中农业大学学报，2025，44（4）：143‐149.

YU Zhen,XU Yurong,WU Zhekuan,et al.Isolation and identification of α-amylase producing bacteria and enzymatic characteristics of Ba8a protein[J].Journal of Huazhong Agricultural University,2025,44(04):143-149.
余振，徐玉荣，吴哲宽，等.α-淀粉酶产生菌的筛选鉴定及Ba8a蛋白的酶学特性［J］.华中农业大学学报，2025，44（4）：143‐149. DOI： 10.13300/j.cnki.hnlkxb.2025.04.014.

YU Zhen,XU Yurong,WU Zhekuan,et al.Isolation and identification of α-amylase producing bacteria and enzymatic characteristics of Ba8a protein[J].Journal of Huazhong Agricultural University,2025,44(04):143-149. DOI： 10.13300/j.cnki.hnlkxb.2025.04.014.

摘要

为解决农作物秸秆降解的问题，采用选择性培养基从牛胃秸秆发酵物中筛选获得1株高产α-淀粉酶的菌株，命名为SCUEC8菌株。通过对SCUEC8菌株进行形态学观察和生理生化特性分析，根据16S rDNA 序列构建系统进化树，鉴定该菌为枯草芽孢杆菌。以SCUEC8菌株的总DNA为模板扩增α-淀粉酶基因

ba8a

，构建pET28a（+）-

ba8a

重组质粒，转化大肠杆菌BL21菌株进行异源表达，进一步研究Ba8a蛋白酶学特性。结果显示，

ba8a

基因大小为1 434 bp，Ba8a蛋白分子质量为52.9 ku ；在pH 5.0、反应温度 45 ℃条件下，Ba8a蛋白相对酶活性较高；Mn

对Ba8a蛋白酶活性具有较高促进作用，Cu

对Ba8a蛋白酶活性具有较高抑制作用；在pH 6.0、反应温度0 ℃条件下，Ba8a蛋白较稳定。结果表明，枯草芽孢杆菌SCUEC8菌株具有α-淀粉酶活性，有助于提高农作物秸秆的降解率。

Abstract

A high-yield strain of α-amylase， named as SCUEC8 strain， was isolated from fermented straw in bovine stomach with selective culture medium to solve the problem of decomposing crop straw. The strain isolated was identified as

Bacillus subtilis

based on the morphological observation and physiological and biochemical characteristics combined with comparative analysis of 16S rDNA sequence. The genomic DNA of SCUEC8 strain was used as template to amplify the

ba8a

gene of α-amylase by PCR. The recombinant plasmid pET28a （+）-

ba8a

was constructed and transformed into the

E. coli

BL21 strain for heterologous expression to further analyze the enzymatic chara

cteristics of Ba8a protein expressed. The results showed that the length of

ba8a

gene was 1 434 bp， and the molecular weight of the Ba8a protein was 52.9 ku. The relative enzyme activity of Ba8a protein was relatively high with pH 5.0 and a reaction temperature of 45 ℃. Mn

had a higher effect on promoting the enzyme activity of Ba8a protein， while Cu

had a higher effect on inhibiting the enzyme activity of Ba8a protein. Ba8a protein was relatively stable at pH 6.0 and temperature 0 ℃. It is indicated that the

Bacillus subtilis

SCUEC8 strain has the activity of α-amylase， which helps to improve the degradation rate of crop straw.

关键词

Keywords

references

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