Fluoroaluminum and fluoroberyllium nucleoside diphosphate complexes as probes of the enzymatic mechanism of the mitochondrial F1-ATPase.
The mechanism by which fluoride and aluminum or beryllium in combination with ADP inhibit beef heart mitochondrial F1-ATPase was investigated. The kinetics of inhibition depended on the nature of the anion present in the F1-ATPase assay medium. Inhibition required the presence of Mg2+ and developed more rapidly with sulfite and sulfate than with chloride, i.e., with anions which activate F1-ATPase activity. The ADP-fluorometal complexes were bound quasi-irreversibly to F1, and each mole of the inhibitory nucleotide-fluorometal complex was tightly associated with 1 mol of Mg2+. One mole of nucleotide-fluorometal complex was able to inhibit the activity of 1 mol of catalytic site in F1. Direct measurements of bound fluoride, aluminum, beryllium, and ADP indicated that the F1-bound ADP-fluorometal complexes are of the following types: ADP1A11F4, ADP1Be1F1, ADP1Be1F2, or ADP1Be1F3. Fluoroaluminates or fluoroberyllates are isomorphous to Pi, and the inhibitory nucleotide-fluorometal complexes mimicked transient intermediates of nucleotides that appeared in the course of ATP hydrolysis. On the other hand, each mole of fully inhibited F1, retained 2 mol of inhibitory complexes. The same stoichiometry was observed when ADP was replaced by GDP, a nucleotide which, unlike ADP, binds only to the catalytic sites of F1. These results are discussed in terms of a stochastic model in which the three cooperative catalytic sites of F1 function in interactive pairs.
Issartel
J P
JP
Departement de Biologie Moléculaire et Structurale, Centre d'Etudes Nucléaires de Grenoble, France.
Dupuis
A
A
Lunardi
J
J
Vignais
P V
PV
D000244:Adenosine Diphosphate / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D000535:Aluminum / Q000378:metabolism; D000818:Animals; D000838:Anions; D001608:Beryllium / Q000378:metabolism; D002384:Catalysis; D002417:Cattle; D005459:Fluorides / Q000378:metabolism; D007700:Kinetics; D008929:Mitochondria, Heart / Q000201:enzymology*; D015394:Molecular Structure; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D000838:Anions; D000244:Adenosine Diphosphate; D000535:Aluminum; D006180:Proton-Translocating ATPases; D001608:Beryllium; D005459:Fluorides