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Free keywords:
Dipole moment; Fluorescence-detected electric dichroism; Rotational relaxation; ATPase, Na+/K+−
Abstract:
The electric parameters of Na+/K+-ATPase labeled by FITC have been characterized by measurements of the fluorescence-detected electric dichroism. The fluorescence emission was measured with polarizers at the magic angle and the light for excitation was usually polarized parallel to the field vector. The FITC-Na+/K+-ATPase preparations exhibit a negative electric dichroism at field strengths up to about 600 V/cm and a positive dichroism at higher field strengths. Pulse reversal experiments reveal a dominant permanent electric moment at low electric field strengths and an increasing contribution from an induced electric moment at higher field strengths. The dichroism rise curves and the transients upon pulse reversal show two relaxation processes with opposite amplitudes, whereas the dichroism decay curves in most cases can be represented by single exponentials at a reasonable accuracy. The amplitude A2 associated with the slower of the rise processes is dominant at low field strengths and also approaches saturation already at low field strengths. The dependence of A2 on the electric field strength is consistent with the orientation function for permanent dipoles and cannot be represented by the orientation function for induced dipoles. The fitted permanent dipole moment is in the range of 3.5 · 10−24 Cm [1 · 106 D] and shows only a relatively small decrease with increasing ionic strength. The stationary values of the electric dichroism up to field strengths E ≤ 800 V/cm can be represented with high accuracy by an orientation function for disk-shaped particles with a permanent moment along the particle symmetry axis and an induced moment along the semi-major axis. The permanent electric moment determined according to this function is consistent with the one obtained from the amplitudes A2. In summary, our measurements indicate that Na+/K+-ATPase is associated with a large permanent electric moment directed perpendicular to the membrane plane. The dipole moment per ATPase monomer unit is estimated to be 1.4 · 10−27Cm [430 ± 50 D].
