Energy transduction in Escherichia coli. The role of the Mg2+ATPase.
Inverted membrane vesicles from strain 7, a wild type Escherichia coli K12 strain, actively transport calcium with energy supplied either by respiration or by ATP. These vesicles also have energy-linked quenching of quinacrine fluorescence. Membranes of strain 7, depleted of Mg2+ATPase by EDTA treatment, lack both activities. Membrane vesicles from strain NR70, a mutant lacking the Mg2+ATPase, show neither calcium transport nor energy-linked fluorescence quenching. Neither EDTA treatment nor genetic loss of the Mg2+atpase causes a reduction in respiration. Purified Mg2+ATPase from strain 7 can bind to EDTA-treated membrane vesicles from either strain 7 or NR70. This binding restored both calcium transport and fluorescence quenching, driven either by respiration or by ATP. Dicyclohexylcarbodiimide treatment mimics the effect of the Mg2+ATPase in the case of respiration-driven reactions. Treatment with EDTA, while not essential for the binding of the Mg2+ATPase to membrane vesicles of NR70, produced better restoration of both activities. The rate of restoration of fluorescence quenching showed a time lag which may indicate that binding of the Mg2+ATPase is a relatively slow process. Antiserum prepared against the Mg2+ATPase inhibited the quenching of quinacrine fluorescence when driven by ATP but not when driven by respiration. Addition of antiserum prior to addition of Mg2+ATPase prevented the restoration of fluorescence quenching, whether driven by respiration or ATP. These results clearly show that MG2+ATPase has an important role not only in catalyzing ATP synthesis and hydrolysis but also in maintaining the energized membrane state.
D000251:Adenosine Triphosphatases / Q000302:isolation & purification / Q000378:metabolism*; D001693:Biological Transport, Active; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D004579:Electron Transport; D004735:Energy Transfer; D004789:Enzyme Activation / Q000187:drug effects; D004926:Escherichia coli / Q000378:metabolism*; D008274:Magnesium / Q000494:pharmacology*; D010101:Oxygen Consumption / Q000187:drug effects; D011796:Quinacrine; D013050:Spectrometry, Fluorescence
D004024:Dicyclohexylcarbodiimide; D000251:Adenosine Triphosphatases; D011796:Quinacrine; D008274:Magnesium; D002118:Calcium