Avtor/Urednik		Gutierrez-Aguirre, Ion; Barlič, Ariana; Podlesek, Zdravko; Maček, Peter; Anderluh, Gregor; Gonzales-Manas, Juan
Naslov		Membrane insertion of the N-terminal alpha-helix of equinatoxin II, a sea anemone cytolytic toxin
Tip		članek
Vir		Biochem J
Vol. in št.		Letnik 384
Leto izdaje		2004
Obseg		str. 421-8
Jezik		eng
Abstrakt		Equinatoxin II (Eqt-II) is a member of the actinoporins, a unique family of cytotoxins comprising 20 kDa pore-fonning proteins isolated crom sea anemones. Actinoporins bind prefereutially to lipid membranes containing sphingomyelin, and create cation-selective pores by oligomerization of three to four monomers. Previous studies have shown that regions of Eql-II crucial for its cytolytic mechanism are an exposed aromatic cluster and the N-tenninal region containing an amphipathic alpha-helix. In the present study, we have investigated the transfer of the N-terminal alpha-helix into the lipid membrane by Ihe use of three mutants containing an additional tryptophan residue in different positions within Ihe amphipathic alpha-helix (Ile18 > Trp, Val22-->Trp and Ala25-->Trp). The interaction of the mutants with different model systems, such as lipid monolayers, erythrocytes and ghost membranes, was extensively characterized.Intrinsic fluorescence measurements and the use of vesicles containing brominated phospholipids indicated a deep localization of the N-terminal amphipathic helix in the lipid bilayer, except for the case of Val22-->Trp. This mutant is stabilized in a state immediately prior to final pore formation. The introduction of additional tryptophan residues in the sequence of Eqt-II has proved to be a suitable approach to monitor the new environments that surround defined regions of the molecule upon membrane interaction.
Deskriptorji		SEA ANEMONES CNIDARIAN VENOMS PORINS CHEMOTACTIC FACTORS MEMBRANE LIPIDS MUTATION LIPID BILAYERS CLONING, MOLECULAR TRYPTOPHAN SPECTROMETRY, FLUORESCENCE