Salt-Induced Oligomerization of Partially Folded Intermediates of Equinatoxin II
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- 作者：Nata
//pubs.acs.org/images/entities/scaron.gif" border="0">a Poklar Ulrih ; Gregor Anderluh ; Peter Ma
- 刊名：Biochemistry
- 出版年：2004
- 出版时间：July 27, 2004
- 年：2004
- 卷：43
- 期：29
- 页码：9536 - 9545
- 全文大小：321K
- 年卷期：v.43,no.29(July 27, 2004)
- ISSN：1520-4995
文摘
Equinatoxin II (EqTxII) is a cytolytic, water-soluble protein which binds to and forms cation-selective pores in lipid membranes. To characterize the native and denatured states of EqTxII and toelucidate the biological role of its oligomers, we have studied salt-dependent heat-induced conformationaltransitions of EqTxII. To this end, we have employed a variety of experimental techniques, includingdifferential scanning calorimetry, circular dichroism and light absorption spectroscopy, ultrasonicvelocimetry, electron microscopy, PAGE, and a hemolytic activity assay. This experimental combinationhas enabled us to monitor and structurally and thermodynamically characterize temperature-inducedconformational transitions and oligomerization of EqTxII at different concentrations of NaCl. At pH 3.0and 25 
C, EqTxII retains its native conformation and remains hemolytically active over a broad rangeof NaCl concentrations. However, an increase in the salt concentration results in a diminution of thethermal stability of EqTxII. Specifically, the calorimetrically determined denaturation temperature, Td,and enthalpy, 
Hcal, of the toxin decrease with an increase in the salt concentration. Our CD data suggestthat the heat-induced denatured state of EqTxII lacks rigid tertiary structure while exhibiting well-definedsecondary structure. The amount of the induced, non-native secondary structure of EqTxII depends onthe solution ionic strength, temperature, time of incubation at an elevated temperature, and proteinconcentration. Our combined results suggest that, in the presence of salt, an increase in temperature resultsin formation of the partially unfolded state of the toxin that oligomerizes and forms biologically inactive,water-soluble aggregates.
C, EqTxII retains its native conformation and remains hemolytically active over a broad rangeof NaCl concentrations. However, an increase in the salt concentration results in a diminution of thethermal stability of EqTxII. Specifically, the calorimetrically determined denaturation temperature, Td,and enthalpy, Hcal, of the toxin decrease with an increase in the salt concentration. Our CD data suggestthat the heat-induced denatured state of EqTxII lacks rigid tertiary structure while exhibiting well-definedsecondary structure. The amount of the induced, non-native secondary structure of EqTxII depends onthe solution ionic strength, temperature, time of incubation at an elevated temperature, and proteinconcentration. Our combined results suggest that, in the presence of salt, an increase in temperature resultsin formation of the partially unfolded state of the toxin that oligomerizes and forms biologically inactive,water-soluble aggregates.