Localization of ortho-nitrophenol within the lipid bilayer

TURINA A. DEL V.; SANCHEZ J.M.; PERILLO M.A.

Hotel 13 de Julio - Mar del Plata, Buenos Aires

Congreso; Congreso Conjunto de Sociedades Biomedicas; 2004

ONP is the product of a b-galactosidase reaction catalyzed. Here we studied the localization of o-nitrophenol (ONP) within model membrane. This is relevant to understand the modulation of b-Gal through its binding to lipid-water interfaces (Sanchez & Perillo, Biophys.Chem.99,281,2002). We measured: 1) the ability of ONP to quench the fluorescence emitted by DPH and TMA-DPH, which localize at different membrane depths, and 2) we studied the acid-base equilibrium of ONP in the presence and in the absence of PC MLVs and Triton X-100 (both electrostatically neutral interfaces), and compared these values with those obtained in solutions of dioxane of known dielectric constants (García & Perillo, BBA,1324,76,1997). ONP quenched TMA-DPH more efficiently than DPH. (a lower concentration of quencher was necessary to reduce the initial fluorescence to a half). The Stern-Volmer plots corresponding to DPH showed a growing exponential behavior which was explained by an increase in the probability of quenching at high quencher concentrations. The DpK of ONP (pKwater - pKTriton= 7.21–7.34= -0.13, nearly zero) was interpolated in a curve of DpK vs the values of dielectric constant of the solutions of 20-80 %P/V dioxane used as solvents. The dielectric constant (DC) value DC=60 obtained reflected the polarity of the bilayers region where the ONP acid-base equilibrium was taking place and indicated a localization of ONP within the polar head group region of the membrane. With PC MLVs the DpK fell outside the range of the experimental curve indicating a contribution of a dipolar potential (60 mV). Our results suggest that ONP (a weak hydrophobic substance) concentrates in the membrane respect to the aqueous phase trough dipolar interactions with phospholipids polar head groups.