Phospholipase A₂ complexes with gadolinium(III) and interaction of the enzyme-metal ion complex with monomeric and micellar alkylphosphorylcholines. Water proton nuclear magnetic relaxation studies

Gadolinium (III) binds competitively with calcium(II) to porcine pancreatic phospholipase A2 (EC 3.1.1.4) and its zymogen. The enzyme-Gd3+ complex exhibits 4% of the hydrolytic activity of the corresponding Ca2+ complex toward a dispersion of dioctanoyllecithin. Dissociation constants for the Gd3+ complex of enzyme and proenzyme were evaluated from water proton relaxation rate (PRR) titrations. At pH 5.8, the dissociation constants for the Gd3+ complexes of enzyme and zymogen are 0.50 and 0.18 mM, respectively. Dissociation constants for the complexes of enzyme with Ca2+, Eu3+, and Tb3+ were evaluated in PRR titrations by competition of these cations with Gd3+ binding. PRR enhancement factors for the Gd3+ complexes of enzyme and proenzyme are 16.4 and 5.8, respectively, at 22 degrees C and 24.3 MHz. Binding of a homologous series of n-alkylphosphorylcholines to the enzyme-Gd3+ complex was investigated through the influence of monomeric and micellar forms of these amphiphiles on the PRR enhancement factor for the enzyme-bound Gd3+. Separate monomer and micelle binding regions were observed in titrations using n-alkylphosphorylcholines with critical micelle concentrations ranging from 15 muM to 13 mM. In every case, the enhancement factors for the enzyme-Gd3+ complexes were significantly greater than that for the tenary complex, enzyme-Gd3+ -monomer. Morever, a synergism was observed in the binding of Gd3+ and micelles to the enzyme. The magnitudes of the PRR enhancement factors for the enzyme-Gd3+ complexes with micelles of n-alkylphosphorylcholines indicate that the bound Gd3+ is freely accessible to the bulk solvent. These results suggest a model for the enzyme-micelle complex in which the active site is spatially removed from the enzyme-micelle interface.