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R. Dean Astumian, PhD
Univ. of Chicago, Surgery/MC 6035, 5841 S Maryland Ave., Chicago, IL
60637
Phone: 312-702-6302; FAX: 312-702-5909
The experiments will be carried out at many different conditions in order to test postulated mechanisms for the field-protein interactions. Some of these include electro-conformational coupling, effects of electric polarization of the double layer, and field inducted changes in ionic concentration in the double layer. We have derived a general expression for the field and amplitude dependence of the response to a weak oscillating field. This can be described as a sum of Lorenzian curves, where the amplitudes depend on the square of the applied field strength. The dependence of these amplitudes on the experimental conditions is different for different mechanisms. Thus, by measuring these amplitudes at different conditions we can distinguish between different mechanisms.
The enzyme used in our study will be the H+ATPase-from yeast because it is readily available in purified form from our collaborator, and because much evidence suggests a strong membrane potential dependence for this enzyme. Thus, there is a very good likelihood of experimental success. In initial experiments we will use enzyme in plasma membranes from fractions with very high specific activity. Later, we will use purified enzyme reconstituted in phospholipid bilayer vesicles.