Interaction of the P-type cardiotoxin with phospholipid membranes.
The cardiotoxin (cytotoxin II, or CTII) isolated from cobra snake (Naja oxiana) venom is a 60-residue basic membrane-active protein featuring three-finger beta sheet fold. To assess possible modes of CTII/membrane interaction 31P- and 1H-NMR spectroscopy was used to study binding of the toxin and its effect onto multilamellar vesicles (MLV) composed of either zwitterionic or anionic phospholipid, dipalmitoylglycerophosphocholine (Pam2Gro-PCho) or dipalmitoylglycerophosphoglycerol (Pam2Gro-PGro), respectively. The analysis of 1H-NMR linewidths of the toxin and 31P-NMR spectral lineshapes of the phospholipid as a function of temperature, lipid-to-protein ratios, and pH values showed that at least three distinct modes of CTII interaction with membranes exist: (a) nonpenetrating mode; in the gel state of the negatively charged MLV the toxin is bound to the surface electrostatically; the binding to Pam2Gro-PCho membranes was not observed; (b) penetrating mode; hydrophobic interactions develop due to penetration of the toxin into Pam2Gro-PGro membranes in the liquid-crystalline state; it is presumed that in this mode CTII is located at the membrane/water interface deepening the side-chains of hydrophobic residues at the tips of the loops 1-3 down to the boundary between the glycerol and acyl regions of the bilayer; (c) the penetrating mode gives way to isotropic phase, stoichiometrically well-defined CTII/phospholipid complexes at CTII/lipid ratio exceeding a threshold value which was found to depend at physiological pH values upon ionization of the imidazole ring of His31. Biological implications of the observed modes of the toxin-membrane interactions are discussed.
Dubovskii
Peter V
PV
Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russian Federation.
Lesovoy
Dmitry M
DM
Dubinnyi
Maxim A
MA
Utkin
Yuri N
YN
Arseniev
Alexander S
AS
D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D004179:Cobra Cardiotoxin Proteins / Q000737:chemistry / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D015394:Molecular Structure; D019906:Nuclear Magnetic Resonance, Biomolecular; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D013696:Temperature
D004179:Cobra Cardiotoxin Proteins; D010743:Phospholipids