title: Phospholipases a2 from Viperidae snakes: Differences in membranotropic activity between enzymatically active toxin and its inactive isoforms.
abstract: We describe the interaction of various phospholipases A2 (PLA2) from snake venoms of the family Viperidae (Macrovipera lebetina obtusa, Vipera ursinii renardi, Bothrops asper) with giant unilamellar vesicles (GUVs) composed of natural brain phospholipids mixture, visualized through fluorescence microscopy. The membrane fluorescent probes 8-anilino-1-naphthalenesulfonicacid (ANS), LAUDRAN and PRODAN were used to assess the state of the membrane and specifically mark the lipid packing and membrane fluidity. Our results have shown that the three PLA2s which contain either of aspartic acid, serine, or lysine residues at position 49 in the catalytic center, have different effects on the vesicles. The PLA2 with aspartic acid at this position causes the oval deformation of the vesicles, while serine and lysine-containing enzymes lead to an appreciable increase of fluorescence intensity in the vesicles membrane, wherein the shape and dimensions of GUVs have not changed, but in this case GUV aggregation occurs. LAURDAN and PRODAN detect the extent of water penetration into the bilayer surface. We calculated generalized polarization function (GP), showing that for all cases (D49 PLA2, S49 PLA2 and K49 PLA2) both LAUDRAN and PRODAN GP values decrease. A higher LAURDAN GP is indicative of low water penetration in the lipid bilayer in case of K49 PLA2 compared with D49 PLA2, whereas the PRODAN mainly gives information when lipid is in liquid crystalline phase.
authors.affiliation: Orbeli Institute of Physiology, Orbely str. 22, 0019 Yerevan, Armenia.
authors.forename: Narine A
authors.initials: NA
authors.lastname: Ghazaryan
authors.affiliation: Orbeli Institute of Physiology, Orbely str. 22, 0019 Yerevan, Armenia.
authors.forename: Lusine
authors.initials: L
authors.lastname: Ghulikyan
authors.affiliation: Orbeli Institute of Physiology, Orbely str. 22, 0019 Yerevan, Armenia.
authors.forename: Arsen
authors.initials: A
authors.lastname: Kishmiryan
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, GSP-7, Ul. Miklukho-Maklaya, 16/10, 117997 Moscow, Russian Federation.
authors.forename: Tatyana V
authors.initials: TV
authors.lastname: Andreeva
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, GSP-7, Ul. Miklukho-Maklaya, 16/10, 117997 Moscow, Russian Federation.
authors.forename: Yuri N
authors.initials: YN
authors.lastname: Utkin
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry of the Russian Academy of Sciences, GSP-7, Ul. Miklukho-Maklaya, 16/10, 117997 Moscow, Russian Federation.
authors.forename: Victor I
authors.initials: VI
authors.lastname: Tsetlin
authors.affiliation: Instituto Clodomiro Picado, Universidad de Costa Rica, 11501 San José, Costa Rica.
authors.forename: Bruno
authors.initials: B
authors.lastname: Lomonte
authors.affiliation: Orbeli Institute of Physiology, Orbely str. 22, 0019 Yerevan, Armenia. Electronic address: taipan@ysu.am.
authors.forename: Naira M
authors.initials: NM
authors.lastname: Ayvazyan
mesh_terms: D015081:2-Naphthylamine / Q000031:analogs & derivatives; D019943:Amino Acid Substitution; D000817:Anilino Naphthalenesulfonates; D000818:Animals; D001224:Aspartic Acid / Q000737:chemistry; D001692:Biological Transport; D001923:Brain Chemistry; D020134:Catalytic Domain; D005456:Fluorescent Dyes; D007848:Laurates; D008051:Lipid Bilayers / Q000737:chemistry*; D008239:Lysine / Q000737:chemistry; D008297:Male; D008560:Membrane Fluidity; D054467:Phospholipases A2 / Q000737:chemistry* / Q000302:isolation & purification; D051381:Rats; D030162:Reptilian Proteins / Q000737:chemistry* / Q000302:isolation & purification; D012694:Serine / Q000737:chemistry; D012910:Snake Venoms / Q000737:chemistry* / Q000201:enzymology; D013329:Structure-Activity Relationship; D053835:Unilamellar Liposomes / Q000737:chemistry*; D017819:Viperidae / Q000378:metabolism; D014867:Water / Q000737:chemistry
chemical_list: C515594:8-anilino-1-naphthalenesulfonic acid; D000817:Anilino Naphthalenesulfonates; D005456:Fluorescent Dyes; D007848:Laurates; D008051:Lipid Bilayers; D030162:Reptilian Proteins; D012910:Snake Venoms; D053835:Unilamellar Liposomes; D014867:Water; D001224:Aspartic Acid; D012694:Serine; C038517:prodan; D015081:2-Naphthylamine; D054467:Phospholipases A2; D008239:Lysine; C065580:laurdan
keywords: Fluorescent probes; GUV; LAURDAN; PRODAN; Phospholipase A2; Snake venom