纤维素降解菌I2的分离及相关降解基因功能验证
Isolation of cellulose-degrading bacterium I2 and functional verification of related degradation genes
孙君珂 1张婉莹 2邓若竹 2王娟 2牛秋红 2胡凡 3柯涛2
作者信息
- 1. 南阳师范学院生命科学与农业工程学院,河南省生物质资源化高校工程中心,河南南阳,473000;自然资源部第三海洋研究所,福建厦门,361000
- 2. 南阳师范学院生命科学与农业工程学院,河南省生物质资源化高校工程中心,河南南阳,473000
- 3. 自然资源部第三海洋研究所,福建厦门,361000
- 折叠
摘要
纤维素是地球上数量最大的可再生能源物质,利用微生物或其产生的酶可将纤维素水解成葡萄糖.筛选高效纤维素降解菌和纤维素酶一直是生物能源等领域研究的热点.该研究筛选分离了 1株具有降解纤维素能力的短小芽孢杆菌I2,通过基因组测序,分析其参与纤维素降解的基因,并对纤维素酶的功能进行验证.I2分离自河南南阳宝天曼自然保护区落叶及长期堆积小麦秸秆的堆体等样本富集后的混合菌系,具有降解滤纸的能力,48 h内可以降解大部分滤纸.基因组组装分析显示,I2的基因组大小为4.2 Mbp,含有4 635个基因.利用CAZy碳水化合物数据库注释和酶活性测定实验,对数据库检测到的182个基因进行分析,明确I2中4个基因在大肠杆菌表达后具有纤维素酶活性.其中,G2269纤维素内切酶活性最高,在pH 5.0,温度55℃条件下,酶活力可达49.02 U/mL.金属离子Ca2+、Zn2+对于G2269的活性有明显提高作用,Mn2+和Cu2+对G2269活性抑制比较明显,Mg2+、Co2+、K+、Ni2+、NH4+对于G2269的活性无显著影响.此外,G2269还具有良好热稳定性和酸碱稳定性,在65 ℃保持30 min后仍保留70％的活力,在食品加工和饲料生产等领域显示出较好的应用潜力.
Abstract
Cellulose is the most abundant renewable energy material on the Earth,and the exploration of highly efficient cellulose-de-grading bacteria and cellulose enzymes has been a focal point of research.In this study,a cellulose-degrading Bacillus strain,designated as I2,was isolated and the functionality of its cellulose genes was validated.I2 was isolated from the accumulated wheat straw in the Bao-tianman Nature Reserve in Nanyang,Henan Province,with a genome size of 4.2 Mbp containing 4 635 genes,capable of degrading a sig-nificant portion of filter paper within 48 hours.Through predictions from the Carbohydrate-Active enZymes(CAZy)carbohydrate database and enzyme activity assays,182 genes detected in the database were analyzed,andit was established that four proteins in I2 exhibit cellu-lose enzyme activity.Among them,G2269 displayed the highest cellulase endoglucanase activity,reaching 49.02 U/mL under pH 5.0 and 55 ℃ conditions.Metal ions Ca2+and Zn2+had a significant enhancing effect on the activity of G2269,while Mn2+and Cu2+signifi-cantly inhibited the activity of G2269.Mg2+,Co2+,K+,Ni2+,and NH4+had no significant effect on the activity of G2269.Further-more,G2269 demonstrated good thermal stability and acid-alkali stability,retaining 70％of its activity after 30 minutes at 65 ℃,presen-ting promising application prospects,particularly in the fields of food processing and feed production.
关键词
纤维素降解菌/基因克隆/原核表达/基因组测序Key words
cellulose degrading bacteria/gene cloning/prokaryotic expression/genome sequencing引用本文复制引用
出版年
2025
国家科技期刊平台
NSTL
维普
万方数据