Enhanced antibacterial activity of an attacin-coleoptericin hybrid protein fused with a helical linker
Enhanced antibacterial activity of an attacin-coleoptericin hybrid protein fused with a helical linker
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DOI：
10.1007/s11033-012-2472-4
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发表时间：
2013-06-01
影响因子：
2.8
通讯作者：
Cho, Saeyoull
中科院分区：
文献类型：
--
作者：
Lee, Minsup;Bang, Kyeongrin;Cho, Saeyoull
此前，我们从甜菜夜蛾 (Spodoptera exigua) 中分离并鉴定了 attacin，并从 Protaetia brevitarsis seulensis 中分离出鞘翅素样蛋白。在这项研究中，我们融合了这两个编码抗菌蛋白的基因，以获得具有增强抗菌活性的杂合蛋白。为了融合两种抗菌蛋白，我们采用了螺旋和非螺旋接头序列，它们在蛋白质中充当域间接头。我们使用 Gly-Gly-Gly-Gly-Ser 肽作为非螺旋接头。使用该接头产生的杂合蛋白对大肠杆菌、枯草芽孢杆菌、荚膜伯克霍尔德氏菌、波纹假单胞菌和鼠疫欧文氏菌的抗菌活性低于两种亲本抗菌蛋白中的任何一种。此外，杂合蛋白的MIC值为23.1μM，表明对大肠杆菌的活性较差。当我们使用三个 Glu-Ala-Ala-Ala-Lys (EAAAK) 肽序列作为螺旋接头来融合两种蛋白质时，所得杂合蛋白比亲本抗菌蛋白具有更高的抗菌活性。特别是，这种杂合蛋白对 P. corrugate 具有很强的抗菌活性。这些结果表明EAAAK基序可用于有效分离两种抗菌蛋白并产生比任一亲本蛋白具有更强抗菌活性的杂合蛋白。
Previously, we isolated and characterized attacin from Spodoptera exigua and a coleoptericin-like protein from Protaetia brevitarsis seulensis. In this study, we fused these two genes encoding antimicrobial proteins to obtain a hybrid protein with enhanced antimicrobial activity. To fuse the two antimicrobial proteins, we employed helical and non-helical linker sequences that function as inter-domain linkers in proteins. We used the Gly-Gly-Gly-Gly-Ser peptide as a non-helical linker. The hybrid protein produced using this linker showed less antimicrobial activity against Escherichia coli, Bacillus subtilis, Burkholderia glumae, Pseudomonas corrugate, and Erwinia rhapontici than either of the two parental antimicrobial proteins. In addition, the MIC value of the hybrid protein was 23.1 mu M, which indicates poor activity against E. coli. When we used three Glu-Ala-Ala-Ala-Lys (EAAAK) peptide sequences as a helical linker to fuse the two proteins, the resultant hybrid protein had much higher antimicrobial activity than the parental antimicrobial proteins. In particular, this hybrid protein had strong antimicrobial activity against P. corrugate. These results indicate that the EAAAK motif can be used to effectively separate two antimicrobial proteins and produce a hybrid protein with more antimicrobial activity than either of the parent proteins.