In this paper, methods for measuring the viscosity and surface tension of low-volume liquid samples are proposed. Viscosity is measured by recording the motion of a liquid driven by surface tension through a horizontal glass capillary tube. Surface tension is determined by connecting a container of a constant volume to the distal end of the capillary tube and measuring the penetration distance of the liquid entering via the proximal end. Sucrose solutions of various concentrations (0–50 wt%) were measured as samples, and the determined viscosities were confirmed to be consistent with the tabulated values provided in the Handbook of Chemistry. The surface tension was found to be constant within the measurement sensitivity of the device regardless of the sucrose concentration.
Bacteriorhodopsin from Halobacterium halobium was stable and active in high concentrations of guanidine hydrochloride. However the decay of the intermediate with absorption maximum at 407 nm and the uptake of protons from the medium were remarkably delayed. At alkaline pH values, all the molecules of bacteriorhodopsin existed as the 407 nm intermediate in the light, and one proton per bacteriorhodopsin molecule was released.
Abstract— Light‐induced proton release and uptake by acetylated and unmodified bacteriorhodopsin were measured. Bacteriorhodopsin, when illuminated, shows a net proton release at neutral and alkaline pH's, but in acidic pH, it shows an uptake of protons. In the presence of high concentrations of guanidine hydrochloride, light caused only proton release even in acidic pH and the maximum extent of the release was one proton per bacteriorhodopsin molecule around pH 8. Acetylation of bacteriorhodopsin caused no alteration in the absorption spectrum of purple complex (bR 570 ) and M 412 ‐intermediate, but decreased the decay rate of the M 412 ‐intermediate. Light‐induced release of protons was not observed even in neutral pH values, and only the proton uptake was noticed by acetylated purple membrane fragments. In high concentrations of guanidine hydrochloride, no proton uptake or release by illumination was observed. Vesicles were reconstituted from acetylated purple membrane. These vesicles had almost no ability for light‐induced proton transport. The role of amino group(s) in light‐induced proton release and transport through the purple membrane is discussed.
Abstract— The decay time course of intermediate M of bacteriorhodopsin was investigated by flash spectrophotometry. The decay was composed of two exponentials showing the existence of two forms of intermediate absorbing around 410 nm. The two were very different in kinetic character whereas the absorption spectra were almost the same. The relative yield of the two components was a function of the intensity of the exciting flash and the slower component disappeared when the flash intensity was made very small. A model based on the trimeric cluster structure of bacteriorhodopsin is proposed.
Proton translocating ATPase (FoF1) from bovine heart mitochondria was reconstituted into planar phospholipid bilayers, and its electrogenicity was directly demonstrated. The FoF1ATPase was solubilized using 3-[(3-cholamidopropyl)-dimethyl-ammonio]-1-propanesulfonic acid (CHAPS) as a detergent followed by sucrose density gradient centrifugation according to the method originally described by McEnery et al. for rat liver mitochondria (McEnery et al. (1986) J.Biol.Chem. 259, 4642–4651), with minor modifications. The purified ATPase was reconstituted into proteoliposomes and then reconstituted into planar phospholipid bilayers by the modified fusion method (Hirataet al. (1986) J.Biol. Chem. 261,9839–9843). A short-circuit current of up to 0.4 pA was induced by adding ATP, and this current was suppressed by the F1 ATPase inhibitor NaN3 or by a specific mitochondrial Fo inhibitor, oligomycin. The direction of the current corresponded to the flow of positive charges from the F1 side to the Fo side. All these facts clearly demonstrate that the mitochondrial FoF1ATPase was successfully reconstituted into planar phospholipid bilayers, and the current was generated by the ATPase.
