College of Biological Science and Engineering, Fuzhou University, Fuzhou, Fujian 350108, China; College of Food Science, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China; Key Laboratory of Postharvest Biology of Subtropical Special Agricultural Products, Fujian Province University, Fuzhou, Fujian 350002, China.
College of Biological Science and Engineering, Fuzhou University, Fuzhou, Fujian 350108, China.
Food Chem. 2021 Jul 1;349:129130. doi: 10.1016/j.foodchem.2021.129130. Epub 2021 Jan 19.
The antifungal protein MG-3A was isolated from Bacillus amyloliquefaciens MG-3, and was purified and identified. The results showed that antifungal protein MG-3A was likely a serine protease with a molecular weight of ~48 kDa. The serine protease exhibited a broad antifungal spectrum and effectively extended the shelf-life of loquat fruit up to 25 d. The antifungal protein MG-3A showed good stabilities to temperature, pH and protease K. Primers were designed according to the mass spectrum of antifungal protein and the comparison with proteins in the NCBI database. The serine protease gene MG-3A from B. amyloliquefaciens genome was isolated and cloned using PCR. The prokaryotic expression plasmid pET28a-MG-3A was constructed and used to express the antimicrobial protein in vitro. The SDS-PAGE results showed that the recombinant protein expressed in Escherichia coli BL21 (DE3) was highly soluble. Affinity chromatography was used to purify the recombinant protein and its antifungal activity was evaluated.
从解淀粉芽孢杆菌 MG-3 中分离到一种抗真菌蛋白 MG-3A，并对其进行了分离和鉴定。结果表明，抗真菌蛋白 MG-3A 可能是一种分子量约为 48 kDa 的丝氨酸蛋白酶。该丝氨酸蛋白酶具有广谱的抗真菌谱，可有效延长枇杷果实的货架期至 25 天。抗真菌蛋白 MG-3A 对温度、pH 和蛋白酶 K 具有良好的稳定性。根据抗真菌蛋白的质谱和与 NCBI 数据库中蛋白质的比较，设计了引物。利用 PCR 从解淀粉芽孢杆菌基因组中分离并克隆了丝氨酸蛋白酶基因 MG-3A。构建了原核表达质粒 pET28a-MG-3A，并在体外表达了抗菌蛋白。SDS-PAGE 结果表明，重组蛋白在大肠杆菌 BL21（DE3）中表达为高度可溶性。亲和层析用于纯化重组蛋白，并评估其抗真菌活性。
