Efficient production and characterization of the novel and highly active antifungal protein AfpB from Penicillium digitatum.
Efficient production and characterization of the novel and highly active antifungal protein AfpB from Penicillium digitatum.
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DOI：
10.1038/s41598-017-15277-w
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发表时间：
2017-11-07
影响因子：
4.6
通讯作者：
Manzanares P
中科院分区：
文献类型：
--
作者：
Garrigues S;Gandía M;Popa C;Borics A;Marx F;Coca M;Marcos JF;Manzanares P
丝状真菌编码独特的抗真菌蛋白（AFPs），为开发新的抗真菌药物提供了巨大的潜力。真菌被认为对自身的磷酸腺苷免疫，就像青霉菌一样，它是众所周知的磷酸腺苷的生产者。指状青霉基因组只编码一个afp基因（afpB），其对应的蛋白（afpB）属于B类系统发育簇。之前检测AfpB的尝试都没有成功。在本研究中，免疫检测证实了野生型和先前重组组成型指状假单胞菌中不存在AfpB积累。利用基于黄芽孢杆菌的表达盒在指状假单胞菌中实现了AfpB的生物技术生产和分泌，用α-因子信号肽在毕赤酵母中实现了AfpB的生物技术生产和分泌。这两种策略都允许适当的蛋白质折叠，高效的生产和从培养上清中一步纯化AfpB。AfpB对大多数真菌的抑菌活性均高于P. chrysogenum PAF，特别是对青霉菌和指状假单胞菌高度敏感。光谱数据表明，天然折叠不是活性所必需的。AfpB还显示出显著的抗蛋白酶和热降解能力，没有溶血活性，使AfpB成为控制病原菌的有希望的候选者。
Filamentous fungi encode distinct antifungal proteins (AFPs) that offer great potential to develop new antifungals. Fungi are considered immune to their own AFPs as occurs in Penicillium chrysogenum, the producer of the well-known PAF. The Penicillium digitatum genome encodes only one afp gene (afpB), and the corresponding protein (AfpB) belongs to the class B phylogenetic cluster. Previous attempts to detect AfpB were not successful. In this work, immunodetection confirmed the absence of AfpB accumulation in wild type and previous recombinant constitutive P. digitatum strains. Biotechnological production and secretion of AfpB were achieved in P. digitatum with the use of a P. chrysogenum-based expression cassette and in the yeast Pichia pastoris with the α-factor signal peptide. Both strategies allowed proper protein folding, efficient production and single-step purification of AfpB from culture supernatants. AfpB showed antifungal activity higher than the P. chrysogenum PAF against the majority of the fungi tested, especially against Penicillium species and including P. digitatum, which was highly sensitive to the self-AfpB. Spectroscopic data suggest that native folding is not required for activity. AfpB also showed notable ability to withstand protease and thermal degradation and no haemolytic activity, making AfpB a promising candidate for the control of pathogenic fungi.
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影响因子：
3.1
作者：
Hegedues, Nikoletta;Sigl, Claudia;Zadra, Ivo;Pocsi, Istvan;Marx, Florentine
通讯作者：
Marx, Florentine
影响因子：
4.3
作者：
Fizil, Adam;Gaspari, Zoltan;Barna, Terezia;Marx, Florentine;Batta, Gyula
通讯作者：
Batta, Gyula
DOI：
10.1111/j.1742-4658.2009.07011.x
发表时间：
2009-05
期刊：
The FEBS journal
影响因子：
--
作者：
Batta G;Barna T;Gáspári Z;Sándor S;Kövér KE;Binder U;Sarg B;Kaiserer L;Chhillar AK;Eigentler A;Leiter E;Hegedüs N;Pócsi I;Lindner H;Marx F
通讯作者：
Marx F
影响因子：
6
作者：
Hegedues, Nikoletta;Marx, Florentine
通讯作者：
Marx, Florentine
DOI：
10.1016/j.ijfoodmicro.2015.03.029
发表时间：
2015-07-16
影响因子：
5.4
作者：
Delgado, Josue;Acosta, Raquel;Asensio, Miguel A.
通讯作者：
Asensio, Miguel A.