Title: Adenosine monophosphate as the first phosphoryl acceptor in oxidative phosphorylation
Abstract: Abstract It was found that AMP as well as ADP could release the controlled respiration of beef heart mitochondria. Stoichiometric measurements showed that, when AMP was the exogenous phosphoryl (32P) acceptor, the value ot the molar ratio (32Pi incorporated/AMP added) was 1.0; the ratio (μmoles AMP/μatoms extra oxygen absorbed) was 2.5 (substrate, pyruvate plus malate); the corresponding P O ratio was 2.5; the molar ratio (ATP labeled/ATP formed) was 1.0. These ratios were not affected by the addition of ATP to the mitochondria prior to AMP addition. The ATP formed when AMP was the added phosphoryl acceptor was exclusively doubly labeled. This was demonstrated by techniques involving column chromatography and enzymic analyses (with hexokinase and myokinase). However, when ADP was added, the values of the ratios were, respectively: (32Pi incorporated/ADP), 0.48; ( ADP O ), 5.1; ( P O ), 2.5; and (ATP labeled/ATP formed), 0.45. In these circumstances the ATP formed was about 15% doubly labeled and about 30% terminally labeled. When AMP was the added phosphate acceptor, the initial rate of 32Pi incorporation (1–2 seconds) was at least twice that observed when ADP was the added acceptor. These results are interpreted as evidence that AMP is the first exogenous acceptor of the phosphoryl group in oxidative phosphorylation, and that release of respiratory control by ADP is mediated through a myokinase reaction. The relative amounts of adenine nucleotides found in the reaction products varied with time of sampling. When AMP was added as phosphoryl acceptor it was found that, at the point of transition of respiration from state 3 to state 4, the nucleotides formed were equally divided between “AMP” and doubly labeled ATP. However, 3 minutes after the transition, a decrease was demonstrated in the amounts of both “AMP” and ATP formed; and an equivalent formation of labeled ADP was demonstrated. When ADP was the nucleotide added as phosphoryl acceptor, the micleotides formed at the transition point consisted of 25% “AMP,” 30–35% labeled ATP, and 40–45% unlabeled ATP. Three minutes later a redistribution ot micleotides was demonstrated with the formation of ADP. From a consideration of these findings and other supporting data it is postulated that the pathway of phosphorylation of exogenous ADP is as follows: 4 ADP → 2 ATP + 2 AMP (external myokinase) (a) 2 AMP + 2Pi∗ → 2 ADP∗ (oxidative phosphorylation) (b) 2 ADP → AMP + ATP∗∗ (internal myokinase) (c) The apparent failure of the AMP produced in reaction (c) to recycle is explained by the finding that the “AMP” found in the reaction products is not in a form which can act to release respiratory control but is in fact probably an acylated AMP. In the case of the mitochondria used in these studies, this product is not rapidly reconverted to free 5′-AMP. Whether “AMP” is a product of another reaction in the phosphorylation pathway synchronous with reactions (b) and (c) or of a completely independent process is not known.
Publication Year: 1966
Publication Date: 1966-11-01
Language: en
Type: article
Indexed In: ['crossref', 'pubmed']
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Cited By Count: 26
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