COMPARISON BETWEEN THE BINDING OF CA2-AFFINITY SITES OF BACTERIORHODOPSIN( AND MG2+ TO THE 2 HIGH)

Bacteriorhodopsin contains Ca2+ and Mg2+ ions whose removal inhibits i ts proton pump function. The binding constants of Ca2+ to the high-aff inity sites were determined by the use of a calcium ion specific elect rode. The unavailability of magnesium ion specific electrode prevented a similar determination for Mg2+. Th, binding constant of Mg2+ to the binding site of highest affinity is determined by using a calcium ion selective electrode to measure the concentration of free Ca2+ in comp etition with Mg2+ for the binding. The binding constant of Mg2+ to the second high affinity site is determined spectrally. The two high-affi nity binding constants for Mg2+ are compared with those obtained for C a2+ in the absence and the presence of Mg2+. The fact that the presenc e of low concentration of one metal ion does not affect the binding co nstant of the other metal ion to the other binding site supports the a ssumption of the independence of the two high-affinity sites of one an other. The difference in the observed values of the binding constants of the two high-affinity sites for Ca2+ and Mg2+ is qualitatively disc ussed in terms of the enthalpy and entropy changes in the binding equi librium.