Topography Studies on the Membrane Interaction Mechanism of the Eosinophil Cationic Protein
详细信息 查看全文
- 作者：Marc Torrent ; Elisabet Cuyá ; s ; Esther Carreras ; Susanna Navarro ; Olga Ló ; pez ; Alfons de la Maza ; M. Victò ; ria Nogué ; s ; Yana K. Reshetnyak ; Ester Boix
- 刊名：Biochemistry
- 出版年：2007
- 出版时间：January 23, 2007
- 年：2007
- 卷：46
- 期：3
- 页码：720 - 733
- 全文大小：471K
- 年卷期：v.46,no.3(January 23, 2007)
- ISSN：1520-4995
文摘
The eosinophil cationic protein (ECP) is an antipathogen protein involved in the host defensesystem. ECP displays bactericidal and membrane lytic capacities [Carreras et al. (2003) Biochemistry 42,6636-6644]. We have now characterized in detail the protein-membrane interaction process. All observedfluorescent parameters of the wild type and single-tryptophan-containing mutants, as well as the resultsof decomposition analysis of protein fluorescence, suggest that W10 and W35 belong to two distinctspectral classes I and III, respectively. Tryptophan residues were classified and assigned to distinct structuralclasses using statistical approaches based on the analysis of tryptophan microenvironment structuralproperties. W10 belongs to class I and is buried in a relative nonpolar, nonflexible protein environment,while W35 (class III) is fully exposed to free water molecules. Tryptophan solvent exposure and thedepth of the protein insertion in the lipid bilayer were monitored by the degree of protein fluorescencequenching by KI and brominated phospholipids, respectively. Results indicate that W35 partially insertsinto the lipid bilayer, whereas W10 does not. Further analysis by electron microscopy and dynamic lightscattering indicates that ECP can destabilize and trigger lipid vesicle aggregation at a nanomolarconcentration range, corresponding to about 1:1000 protein/lipid ratio. No significant leakage of the vesicleaqueous content takes place below that protein concentration threshold. The data are consistent with amembrane destabilization "carpet-like" mechanism.