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.
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— 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.
Proteoliposome vesicles containing both bacteriorhodopsin of Halobacterium halobium and H+-translocating ATPase [EC 3.6,1.3] of a thermophilic bacterium, PS3, (TF0-F1) were reconstituted by either the dialysis method or the sonication method. Generation of the electrochemical proton gradient (deltamuH+) in these vesicles was measured using 9-aminoacridine for estimation of the chemical (deltapH) component and 8-anilinonaphthalene sulfonate for the electrical (deltaphi) component). In illuminated bacteriorhodopsin-vesicles the deltamuH+ reached 180-190 mV when reconstituted by the dialysis method and 210-220 mV when reconstituted by the sonication method. Vesicles reconstituted from both TF0-F1 and bacteriorhodopsin by the dialysis method generated a deltapH+ of about 200 mV on addition of ATP, while vesicles prepared by the sonication method generated very little deltamuH+, if any. These vesicles generated similar deltamuH+ on illumination to that found in bacteriorhodopsin-vesicles. Using vesicles reconstituted from both TF0-F1 and bacteriorhodopsin by the dialysis method, light dependent ATP synthesis was measured in relation to deltamuH+ formation. It was necessary to generate a deltamuH+ of above 170 mV for demonstration of appreciable formation of ATP and the greater the deltamuH+, the faster the rate of ATP synthesis.
An ATPase was extracted and purified from castor bean endosperm mitochondria. The enzyme is stable at 60°C only in the presence of ATP in the incubation medium. It is less stable at 0°C than at 30°C but is stabilized by ammonium sulfate or glycerol. Activity is dependent on the presence of Mg++, and in the presence of Mg++ is enhanced by 2,4-dinitrophenol, but is not inhibited by oligomycin. The enzyme hydrolyzes ITP in addition to ATP, but ITPase activity is hardly enhanced by 2,4-dinitrophenol. This preparation has many properties in common with the ATPase (coupling factor 1) from beef heart mitochondria.
Journal Article Light-Induced Formation of the 410 nm Intermediate from Reconstituted Bacteriorhodopsin Get access Koki OHNO, Koki OHNO *Department of Physics, Jichi Medical SchoolYakushiji, Tochigi 329-04 Search for other works by this author on: Oxford Academic PubMed Google Scholar Yasuaki TAKEUCHI, Yasuaki TAKEUCHI **Department of Biology, Jichi Medical SchoolYakushiji, Tochigi 329-04 Search for other works by this author on: Oxford Academic PubMed Google Scholar Masasuke YOSHIDA Masasuke YOSHIDA ***Department of Biochemistry, Jichi Medical SchoolYakushiji, Tochigi 329-04 Search for other works by this author on: Oxford Academic PubMed Google Scholar The Journal of Biochemistry, Volume 82, Issue 4, October 1977, Pages 1177–1180, https://doi.org/10.1093/oxfordjournals.jbchem.a131792 Published: 01 October 1977 Article history Received: 15 June 1977 Published: 01 October 1977
