Structure and stability of the snail neurotoxin PV2.

FRASSA,V.; DREON, M.S.; CEOLIN,M.; HERAS, H.

SM Tucuman

Congreso; XLV Reunión SAIByBM,; 2009

XLV Reunión SAIByBM,

PV2 is one of the major egg proteins from the perivitellin fluid of the eggs of the snail Pomacea canaliculata. It is an oligomer of ca.400 KDa with nutritive and neurotoxic properties. Here we characterize PV2 regarding oligomer structure, size and global shape, pH stability and resistance to proteolysis. Fluorescence and electrophoretic assays suggest PV2 is an octamer of 4 identical 98 KDa heterodimers composed of 67 and 31 KDa subunits which are held together by disulfide bridges. Dimers are assembled into native PV2 by non-covalent forces. Proteinase K treatment suggests the 67 KDa subunit is more exposed. Analysis by small angle X-ray scattering (SAXS) showed that PV2 is an anisometric particle of 130 x 44Å, a size coincident with electron microscopy results. The 3D shape model presented is the first for an egg proteinaceous neurotoxin. Native PV2 isoelectric point is 6.2 and its stability against pH, studied by intrinsic tryptophan fluorescence and SAXS, showed that PV2 conformation is extremely stable under a wide pH range (2.0-12.0). PV2 was also resistant to trypsinolysis in vitro even under 0.1 % SDS, though trypsin is able to digest the heat-denatured protein. PV2 resistance to proteolysis by trypsin and stability under a wide pH range suggests a novel mechanism to avoid digestion by predators that allow PV2 to maintain its neurotoxic properties when ingested.