Expression and purification of recombinant puroindoline A protein in Escherichia coli and its antifungal effect against Aspergillus flavus
Expression and purification of recombinant puroindoline A protein in Escherichia coli and its antifungal effect against Aspergillus flavus
复制标题
重组嘌呤吲哚啉A蛋白在大肠杆菌中的表达、纯化及其对黄曲霉的抗真菌作用
DOI：
10.1007/s00253-019-10168-8
复制
发表时间：
2019-11
影响因子：
5
通讯作者：
Yuansen Hu
中科院分区：
文献类型：
--
作者：
Ang Lv;Cuixiang Li;Pingping Tian;Wenjing Yuan;Shuaibing Zhang;Yangyong Lv;Yuansen Hu
黄曲霉是造成农产品采后商品损失的主要原因。本研究在大肠杆菌中表达并纯化了嘌呤吲哚啉A（PINA）蛋白，并研究了PINA蛋白对A。flavus的特征。十二烷基硫酸钠聚丙烯酰胺凝胶电泳显示重组PINA蛋白的分子量约为44 kDa。PINA对A.在马铃薯葡萄糖琼脂培养基上，黄曲霉的最适生长浓度为42.42 μg/mL。流式细胞仪和扫描电镜显示，孢子形态被破坏的PINA曝光;孢子被压低和破碎，表明细胞壁受损。透射电子显微镜和碘化丙啶染色说明细胞内孢子结构的显着变化，表明细胞膜损伤。5，5 '，6，6'-四氯-1，1 '，3，3'-四乙基苯并咪唑羰花青碘化物染色显示线粒体膜电位降低。4 '，6-二脒基-2-苯基吲哚染色可见大的核凝聚和DNA片段化。与细胞壁、细胞膜和孢子萌发相关的基因表达在PINA处理的细胞中发生了显著变化，这说明了PINA对A.通过细胞壁破坏和触发细胞膜、线粒体和DNA损伤导致细胞死亡而引起黄曲霉。小麦PINA蛋白对A.黄曲霉在农业生产中具有潜在的应用价值。
Aspergillus flavus is the main cause of postharvest agricultural commodity loss. In this study, puroindoline A (PINA) protein was expressed in Escherichia coli, purified, and its antifungal properties against A. flavus were characterized. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that the molecular weight of the recombinant PINA protein was approximately 44 kDa. PINA exerted a powerful antifungal effect against A. flavus at 42.42 μg/mL on potato dextrose agar culture medium. Flow cytometry and scanning electron microscopy revealed that the spore morphology was damaged by PINA exposure; spores were depressed and broken, suggesting that the cell wall was impaired. Transmission electron microscopy and propidium iodide staining illustrated significant changes in intracellular spore structure, indicating cell membrane damage. 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine iodide staining indicated decreased mitochondrial membrane potential. Large nuclear condensation and DNA fragmentation were detected by 4',6-diamidino-2-phenylindole staining. The expression of genes related to the cell wall, cell membrane, and spore germination significantly changed in PINA-treated cells; this illustrated the probable mode of PINA action on A. flavus through cell wall destruction and triggered cell membrane, mitochondrial, and DNA damage leading to cell death. The antifungal mechanism of wheat PINA protein on A. flavus has been demonstrated in this study, and has potential application in preventing postharvest loss in the agricultural industry.
登录
查看更多内容
影响因子：
2.8
作者：
J. Ramos;Pilar Contreras;A. Rodríguez-Navarro
通讯作者：
J. Ramos;Pilar Contreras;A. Rodríguez-Navarro
影响因子：
2.9
作者：
Bihan, TL;Blochet, JE;Pezolet, M
通讯作者：
Pezolet, M
DOI：
10.1104/pp.113.1.83
发表时间：
1997-01
期刊：
--
影响因子：
--
作者：
K. Nielsen;John E. Nielsen;Susan Mampusti Madrid;J. Mikkelsen
通讯作者：
K. Nielsen;John E. Nielsen;Susan Mampusti Madrid;J. Mikkelsen
影响因子：
3.5
作者：
Yingjie Miao;Ling Chen;Cheng Wang;Yajuan Wang;Qian Zheng;Chunbao Gao;Guangxiao Yang;G. He
通讯作者：
Yingjie Miao;Ling Chen;Cheng Wang;Yajuan Wang;Qian Zheng;Chunbao Gao;Guangxiao Yang;G. He
影响因子：
11.4
作者：
HAN, ST;NAVARRO, J;ADAMS, TH
通讯作者：
ADAMS, TH