42 research outputs found
Protonation of a novel intermediate P is involved in the M â bR step of the bacteriorhodopsin photocycle
AbstractA novel intermediate (P) of the bacteriorhodopsin (bR) photocycle, appearing between M412 and bR is described. Like bR, intermediate P shows an absorption maximum at 560â570 nm. However, the extinction coefficient of P is somewhat lower than that of bR. Moreover, there are some differences in spectra of bR and P at wavelengths shorter than 450 nm. The P â bR transition correlates with the absorption of H+ from the water medium. The following conditions proved to be favourable for the detection of the new intermediate: a high salt concentration, low light intensity and low temperature (0.5°C). The P â bR transition is strongly decelerated by a small amount of Triton X-100. Illumination of P does not produce M412 before bR is formed. It is assumed that M412 converts to P when the Schiff base is protonated by a proton transferred from a protein protolytic group which participates in the inward H+-conductivity pathway. Reprotonation of this group results in the conversion of P to bR. No more than 1 H+ is transported per bR photocycle
Editorial vol. 3
Direct measurement of the electrogenic activity of purified mitochondrial transhydrogenase has been carried out. To this end, beefâheart transhydrogenase was isolated and reconstituted with phospholipids to form proteoliposomes. The transhydrogenase proteoliposomes were incorporated into a membrane filter impregnated with a decane solution of phospholipids. It is shown that addition of substrates of either the forward (NADPH and NAD+) or the reverse (NADH and NADP+) transhydrogenase reaction gives rise to an electric potential difference across the proteoliposometreated membrane filter. The electric vector depends upon the direction of the reaction. The proteoliposomeâsupplemented compartment charges negatively in the case of the forward reaction and positively in the case of the reverse one. Addition of the reaction products after substrates equalizes the potentials. The transhydrogenaseâtreated membrane filter retains the ability to perform transhydrogenaseâlinked electrogenesis after removal of excess nonâincorporated proteoliposomes. The electric potential difference reaching 20 mV immediately after the transhydrogenase substrate addition, slowly decreases due to accumulation of the reaction products. Such decay is prevented when the mixture is supplemented with the substrateâregenerating and productâutilizing enzymic systems. Under these conditions, a steady continuous electric current of about 10 pA can be observed. Copyright © 1980, Wiley Blackwell. All rights reserve
Reconstitution of Biological Molecular Generators of Electric Current: Transhydrogenase
Direct measurement of the electrogenic activity of purified mitochondrial transhydrogenase has been carried out. To this end, beefâheart transhydrogenase was isolated and reconstituted with phospholipids to form proteoliposomes. The transhydrogenase proteoliposomes were incorporated into a membrane filter impregnated with a decane solution of phospholipids. It is shown that addition of substrates of either the forward (NADPH and NAD+) or the reverse (NADH and NADP+) transhydrogenase reaction gives rise to an electric potential difference across the proteoliposometreated membrane filter. The electric vector depends upon the direction of the reaction. The proteoliposomeâsupplemented compartment charges negatively in the case of the forward reaction and positively in the case of the reverse one. Addition of the reaction products after substrates equalizes the potentials. The transhydrogenaseâtreated membrane filter retains the ability to perform transhydrogenaseâlinked electrogenesis after removal of excess nonâincorporated proteoliposomes. The electric potential difference reaching 20 mV immediately after the transhydrogenase substrate addition, slowly decreases due to accumulation of the reaction products. Such decay is prevented when the mixture is supplemented with the substrateâregenerating and productâutilizing enzymic systems. Under these conditions, a steady continuous electric current of about 10 pA can be observed. Copyright © 1980, Wiley Blackwell. All rights reserve