The Biological Activity of the Prototypic Cyclotide Kalata B1 Is Modulated by the Formation of Multimeric Pores
The Biological Activity of the Prototypic Cyclotide Kalata B1 Is Modulated by the Formation of Multimeric Pores
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DOI：
10.1074/jbc.m109.003384
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发表时间：
2009-07-31
影响因子：
4.8
通讯作者：
Craik, David J.
中科院分区：
文献类型：
--
作者：
Huang, Yen-Hua;Colgrave, Michelle L.;Craik, David J.
环肽是一大类含有胱氨酸结基序的环状微型蛋白。它们在植物中表达为防御相关蛋白，具有杀虫活性。在这里，我们研究它们在膜相互作用和破坏中的作用。 Kalata B1 是一种原型环肽，被发现可诱导自猝灭荧光团（羧基荧光素）从磷脂囊泡中泄漏。 kalata B1 的丙氨酸扫描诱变表明，裂解活性必需的残基聚集在一起，形成生物活性面。 Kalata B1 被隔离在膜表面并显示出与囊泡的缓慢解离。电生理实验表明，与 kalata B1 一起孵育时，脂质体贴片中会诱导产生导电孔。从电流-电压关系计算的电导表明双层贴片中形成的孔的直径为41-47埃。总的来说，这些发现为这个令人着迷的超稳定大环肽家族的生物功能多样性提供了机制解释。
The cyclotides are a large family of circular mini-proteins containing a cystine knot motif. They are expressed in plants as defense-related proteins, with insecticidal activity. Here we investigate their role in membrane interaction and disruption. Kalata B1, a prototypic cyclotide, was found to induce leakage of the self-quenching fluorophore, carboxyfluorescein, from phospholipid vesicles. Alanine-scanning mutagenesis of kalata B1 showed that residues essential for lytic activity are clustered, forming a bioactive face. Kalata B1 was sequestered at the membrane surface and showed slow dissociation from vesicles. Electrophysiological experiments showed that conductive pores were induced in liposome patches on incubation with kalata B1. The conductance calculated from the current-voltage relationship indicated that the diameter of the pores formed in the bilayer patches is 41-47 angstrom. Collectively, the findings provide a mechanistic explanation for the diversity of biological functions ascribed to this fascinating family of ultrastable macrocyclic peptides.