The antifungal peptide CGA-N12 inhibits cell wall synthesis of Candida tropicalis by interacting with KRE9
The antifungal peptide CGA-N12 inhibits cell wall synthesis of Candida tropicalis by interacting with KRE9
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抗真菌肽 CGA-N12 通过与 KRE9 相互作用抑制热带念珠菌细胞壁合成
DOI：
10.1042/bcj20190678
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发表时间：
2020-02-01
影响因子：
4.1
通讯作者：
Fu, Caiwei
中科院分区：
文献类型：
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作者：
Li, Ruifang;Liu, Zhengwei;Fu, Caiwei
CGA-N12是一种来源于嗜铬粒菌素A的抗真菌多肽，通过诱导细胞凋亡，对念珠菌，特别是对热带念珠菌具有特异的拮抗活性。然而，CGA-N12对念珠菌细胞壁的影响尚不清楚。用亲和层析法对具有β-1，6-葡聚糖酶活性的念珠菌蛋白KRE9与CGA-N12结合后进行了筛选。本研究通过研究CGA-N12对KRE9的影响以及CGA-N12与KRE9之间的相互作用来阐明CGA-N12对热带假丝酵母细胞壁合成的影响。通过分析葡萄糖的消耗，考察了CGA-N12对重组KRE9β-1，6-葡聚糖酶活性的影响。结果表明，CGA-N12对KRE9活性有抑制作用。经CGA-N12处理后，热带锥虫的细胞壁结构遭到破坏。用等温滴定量热法分析了CGA-N12与KRE9的相互作用。结果表明，它们之间的相互作用过程是吸热反应，相互作用力主要是疏水作用力，也有少量静电作用力。荧光共振能量转移(FRET)分析表明，CGA-N12与KRE9相互作用的距离约为7~10 nm。因此，我们得出结论，CGA-N12在热带毛虫细胞膜上的靶标是KRE9，CGA-N12与KRE9弱结合的距离约为10 nm，并抑制KRE9的活性。
CGA-N12, an antifungal peptide derived from chromogranin A, has specific antagonistic activity against Candida spp., especially against Candida tropicalis, by inducing cell apoptosis. However, the effect of CGA-N12 on the Candida cell wall is unknown. The Candida protein KRE9, which possesses beta-1,6-glucanase activity, was screened by affinity chromatography after binding to CGA-N12. In this study, the effect of CGA-N12 on KRE9 and the interaction between CGA-N12 and KRE9 was studied to clarify the effect of CGA-N12 on C. tropicalis cell wall synthesis. The effect of CGA-N12 on recombinant KRE9 beta-1,6-glucanase activity was investigated by analyzing the consumption of glucose. The results showed that CGA-N12 inhibited the activity of KRE9. After C. tropicalis was treated with CGA-N12, the structure of the C. tropicalis cell wall was damaged. The interaction between CGA-N12 and KRE9 was analyzed by isothermal titration calorimetry (ITC). The results showed that their interaction process was involved an endothermic reaction, and the interaction force was mainly hydrophobic with a few electrostatic forces. The results of the fluorescence resonance energy transfer (FRET) assay showed that the distance between CGA-N12 and KRE9 was 7 similar to 10 nm during their interaction. Therefore, we concluded that the target of CGA-N12 in the C. tropicalis cell membrane is KRE9, and that CGA-N12 weakly binds to KRE9 within a 7 similar to 10 nm distance and inhibits KRE9 activity.