CHLORIDE-DEPENDENT AND PH-DEPENDENT PROTON TRANSPORT BY BR MUTANT D85N

Photocurrents from purple membrane suspensions of D85N BR mutant adsor bed to planar lipid membranes (BLM) were recorded under yellow (lambda > 515 nm), blue < 360 nm < lambda < 420 nm) and white (lambda > 360 n m) light. The pH dependence of the transient and stationary currents w as studied in the range from 4.5 to 10.5. The outwardly directed stati onary currents In yellow and blue Light indicate the presence of a pro ton pumping activity, dependent on the pH of the sample, in the same d irection as in the wild-type. The inwardly directed currents in white light, due to an inverse proton translocation, in a two-photon process , show a pH dependence as well. The stationary currents in blue and wh ite light are drastically increased in the presence of azide, but not in yellow light. The concentration dependence of the currents on azide indicates binding of azide to the protein. In the presence of 1 M sod ium chloride, the stationary proton currents in yellow light show an i ncrease by a factor of 25 at pH 5.5. On addition of 50 mM azide, the s tationary current in yellow light decreases again, possibly by competi tion between azide and chloride for a common binding site. The observe d transport modes are discussed in the framework of the recently publi shed IST model for ion translocation by retinal proteins [U. Haupts et al., Biochemistry 36 (1997) 2-7]. (C) 1998 Elsevier Science B.V.