The mitochondrial ATPase. Evidence for a single essential tyrosine residue.
1. Evidence is presented which indicates that inactivation of the mitochondrial ATPase from bovine heart by the reagent 4-chloro-7-nitrobenzofurazan results from modification of one tyrosine residue per enzyme molecule. Activity can be restored by a variety of sulphydryl reagents. 2. In sodium dodecyl sulphate, the nitrogenzofurazan group on tyrosine is transfered to newly exposed sulphydryl groups on the enzyme. 3. The rate of transfer of the nitrobenzofurazan moiety from theenzyme to sulphydryl compounds is compared with that for transfer from the model compound N-acetyl-tyrosine-0(7-nitrobenzo-furazan) ethyl ester, the synthesis and properties of which are also described. 4. The ligands ATP and ADP exert a protective effect on the rate of reaction between the mitochondrial ATPase and 4-chloro-7-nitrobenzofurazan. The variation in rate of this reaction with change in pH has also been examined and a pKa of 9.5 estimated for the tyrosine residue. 5. The modification does not prevent substrate binding as judged by changes in the fluorescence of aurovertin, an antibiotic with specific affinity for mitochondiral ATPases. 6. When the ATPase activity of submitochondrial particles is inhibited by 4-chloro-7-nitrobenzo-furazan, there is a parallel decrease in the extent of the energy-linked fluorescence enhancement of 1-anilino-naphthalene-8-sulphonate induced by ATP hydrolysis. Both ATPase activity and the fluorescence enhancement are restored by sluphydryl reagents.
D000227:Adenine Nucleotides / Q000494:pharmacology; D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors; D000818:Animals; D001313:Aurovertins; D002417:Cattle; D055598:Chemical Phenomena; D002621:Chemistry; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008931:Mitochondria, Muscle / Q000201:enzymology*; D009206:Myocardium / Q000201:enzymology* / Q000648:ultrastructure; D010069:Oxadiazoles / Q000494:pharmacology; D011485:Protein Binding; D012967:Sodium Dodecyl Sulfate; D013050:Spectrometry, Fluorescence; D013438:Sulfhydryl Compounds; D013774:Tetranitromethane / Q000494:pharmacology; D014443:Tyrosine / Q000031:analogs & derivatives
D000227:Adenine Nucleotides; D001313:Aurovertins; D010069:Oxadiazoles; D013438:Sulfhydryl Compounds; D012967:Sodium Dodecyl Sulfate; D014443:Tyrosine; D000251:Adenosine Triphosphatases; D013774:Tetranitromethane