Correlation of photochemical cycle, H + release and uptake, and electric events in bacteriorhodopsin

Abstract

The kinetics of 3 photoinduced responses of bacteriorhodopsin have been compared: spectral changes, pH shifts in a suspension of open purple membrane sheets and electric potential generation by the sheets incorporated into a lipid-impregnated collodion film. In the presence of a pH-buffer, the H + release by bacteriorhodops in was shown to correlate with the formation of the M412 intermediate and the microsecond phase of the potential generation. The H +/M412 ratio is equal to 0.7±0.1 if the ionic strength of the solution is high. In the absence of the buffer, the H + release proved to be much slower than spectral and electric responses. The kinetics of H + uptake by bacteriorhodopsin is close to M412 decay and to the electrogeneous millisecond phase in both the presence and absence of the pH buffer. The bacteriorhodopsin-induced proton release phase accounts for about 20%, and the uptake phase for about 80% of the overall potential. This is compatible with the model assuming that the proton start-out point - possibly, the protonated Schiff base connecting lysine 216 with retinal - is closer to the outer rather than the inner (cytoplasmic) surface of the bacterial membrane.