Identification of a Membrane-Bound Prepore Species Clarifies the Lytic Mechanism of Actinoporins

MORANTE K; BELLOMIO A; GIL-CARTÓN D; REDONDO-MORATA L; SOT J; SCHEURING S; VALLE M; GONZALEZ-MAÑAS JM; TSUMOTO K; CAAVEIRO JMM

JOURNAL OF BIOLOGICAL CHEMISTRY

AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

Lugar: Bethesda, Maryland; Año: 2016

0021-9258

Pore-formingtoxins (PFT) are cytolytic proteins belonging to the molecularwarfare apparatus of living organisms. The assembly of the functionaltransmembrane pore requires several intermediate steps ranging from awater-soluble monomeric species to the multimeric ensemble insertedin the cell membrane. The non-lytic oligomeric intermediate known asprepore plays an essential role in the mechanism of insertion of theclass of β-PFT. However, in the class of α-PFT like theactinoporins produced by sea anemones, evidence of membrane-boundprepores is still lacking. We have employed single-particlecryo-electron microscopy (cryo-EM) and atomic force microscopy (AFM)to identify, for the first time, a prepore species of the actinoporinfragaceatoxin C (FraC) bound to lipid vesicles. The size of theprepore coincides that of the functional pore, except for thetransmembrane region, which is absent in the prepore. Biochemicalassays indicated that, in the prepore species, the N-terminus is notinserted in the bilayer but exposed to the aqueous solution. Ourstudy reveals the structure of the prepore complex in actinoporins,and highlights the role of structural intermediates for the formationof cytolytic pores by an α-PFT.