Photobleaching Reveals Heterogeneous Stoichiometry for Equinatoxin II Oligomers

被引：31

作者：

Baker, Matthew A. B. ^{[1
]}

Rojko, Nejc ^{[2
]}

Cronin, Brid ^{[1
]}

Anderluh, Gregor ^{[2
,3
]}

Wallace, Mark I. ^{[1
]}

机构：

[1] Univ Oxford, Chem Res Lab, Oxford OX1 3QZ, England

[2] Univ Ljubljana, Dept Biol, Biotech Fac, Ljubljana 1000, Slovenia

[3] Natl Inst Chem, Lab Mol Biol & Nanobiotechnol, Ljubljana 1000, Slovenia

来源：

CHEMBIOCHEM | 2014年 / 15卷 / 14期

基金：

英国生物技术与生命科学研究理事会; 英国工程与自然科学研究理事会;

关键词：

equinatoxin; fluorescence; membrane proteins; photobleaching; pore-forming toxins; single molecule fluorescence; PORE-FORMING TOXIN; CHOLESTEROL-DEPENDENT CYTOLYSINS; SEA-ANEMONE; STICHOLYSIN-II; MEMBRANE PERMEABILIZATION; LIPID-MEMBRANES; PROTEIN TOXINS; MECHANISM; SPHINGOMYELIN; FLUORESCENCE;

D O I：

10.1002/cbic.201300799

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

070307 [化学生物学]; 071010 [生物化学与分子生物学];

摘要：

Equinatoxin II (EqtII), a sea anemone cytolysin, is known to oligomerize to form pores that spontaneously insert into membranes. Crystallographic and cryo-EM studies of structurally similar cytolysins offer contradictory evidence for pore stoichiometry. Here we used single-molecule photobleaching of fluorescently labeled EqtII to determine the stoichiometry of EqtII oligomers in supported lipid bilayers. A frequency analysis of photobleaching steps revealed a log-normal distribution of stoichiometries with a mean of 3.4 +/- 2.3 standard deviations. Comparison of our experimental data with simulations of fixed stoichiometries supports our observation of a heterogeneous distribution of EqtII oligomerization. These data are consistent with a model of EqtII stoichiometry where pores are on average tetrameric, but with large variation in the number of subunits in individual pores.

引用

页码：2139 / 2145

页数：7

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