Rabbit muscle phosphofructokinase. 1. Activation by affinity labeling approximately two adenosine cyclic 3',5'-monophosphate binding sites per tetramer.
The smallest enzymatically active form of rabbit muscle phosphofructokinase consists of four identical or nearly identical subunits, and each subunit contains one binding site specific for the activating adenine nucleotides cAMP, AMP, and ADP. These activator binding sites on the enzyme have been covalently labeled to various degrees, ranging from an average value of less than one label/tetramer to four labels/tetramer, with the affinity label 5'-[p-(fluorosulfonyl)-benzoyl]adenosine, and the kinetic and regulatory properties of these modified phosphofructokinase preparations have been investigated. The kinetic and regulatory properties of the affinity-labeled phosphofructokinase are essentially identical with those of native enzyme activated by cAMP, and a near maximum activation of the affinity-labeled enzyme is observed in those preparations modified to the extent of two or more groups/tetramer, suggesting that the covalent attachment of approximately two affinity labels/tetramer is necessary and sufficient for full activation of the enzyme. This requirement for approximately two groups/tetramer for full activation of the enzyme has been substantiated by dissociating a solution containing both native enzyme and affinity-labeled enzyme modified to the extent of approximately four groups/tetramer and then allowing the resulting solution of labeled and unlabeled monomers to reassemble into active tetramers and observing that the extent of activation in the solution of reassembled enzyme is greater than that initially observed in the enzyme solution before dissociation.
D000241:Adenosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000244:Adenosine Diphosphate / Q000378:metabolism; D000249:Adenosine Monophosphate / Q000378:metabolism; D000345:Affinity Labels / Q000494:pharmacology*; D000818:Animals; D001665:Binding Sites; D000242:Cyclic AMP / Q000378:metabolism*; D004789:Enzyme Activation; D009132:Muscles / Q000201:enzymology; D010732:Phosphofructokinase-1 / Q000378:metabolism*; D011817:Rabbits
D000345:Affinity Labels; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; C010959:5'-(4-fluorosulfonylbenzoyl)adenosine; D000242:Cyclic AMP; D010732:Phosphofructokinase-1; D000241:Adenosine