pH dependence of adenosine 5'-triphosphate synthesis and hydrolysis catalyzed by reconstituted chloroplast coupling factor.
The purified ATP-synthesizing complex from chloroplasts has been reconstituted into phospholipid vesicles with bacteriorhodopsin by use of octyl glucoside. Phosphorylation rates up to 90 mmol of ATP (mg of protein)-1 min-1 have been achieved. The dependence of the steady-state kinetic parameters on external and internal pH for both synthesis and hydrolysis was determined. The Michaelis constants are independent of the magnitude of the pH gradient at external pH values of 6.6 and 8.0. The dependence of the maximum velocity for ATP synthesis on the external pH is bell shaped at a constant pH gradient with a maximum at about pH 6.7. The variation of the maximum velocity with external pH is not dependent on the magnitude of the pH gradient. At external pH values of 6.6 and 8.0, the maximum velocity for ATP synthesis varies with approximately the 2.3 power of the internal hydrogen ion concentration. The maximum velocity for ATP hydrolysis also is dependent on the external pH, with a maximum at about pH 8.4; however, most of the ATPase activity is not coupled to the proton flux. Both Mg2+ and Mn2+ are good cofactors for ATP synthesis and hydrolysis whereas Ca2+ is completely ineffective for synthesis and only about 10% as effective as Mg2+ and Mn2+ for hydrolysis. The results obtained suggest that ATP synthesis or hydrolysis may be coupled to proton pumping indirectly, as, for example, by conformational changes.
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- Spectrometry, Fluorescence
- Proton-Translocating ATPases
- Phosphorylation
- Liposomes
- Kinetics
- Hydrogen-Ion Concentration
- Halobacterium
- Biochemistry & Molecular Biology
- Bacteriorhodopsins
- Adenosine Triphosphate
Citation
Published In
DOI
ISSN
Publication Date
Volume
Issue
Start / End Page
Location
Related Subject Headings
- Spectrometry, Fluorescence
- Proton-Translocating ATPases
- Phosphorylation
- Liposomes
- Kinetics
- Hydrogen-Ion Concentration
- Halobacterium
- Biochemistry & Molecular Biology
- Bacteriorhodopsins
- Adenosine Triphosphate