The concentration and control of cytoplasmic free inorganic pyrophosphate in rat liver in vivo.

The concentration of cytoplasmic free pyrophosphate was calculated in freeze-clamped livers of rats from the measured concentration of reactants and K(eq.) of the UDP-glucose pyrophosphorylase reaction (UDP-alpha-d-glucose 1-phosphate uridylyltransferase, EC 2.7.7.9). The K(eq.) of the UDP-glucose pyrophosphorylase reaction was redetermined at 38 degrees C, pH7.0, I=0.25mol/l and free [Mg(2+)]=1mm, and was 4.55 in the direction of glucose 1-phosphate formation. The activity of UDP-glucose pyrophosphorylase in rat liver was between 46 and 58mumol of glucose 1-phosphate formed/min per g fresh wt. in the four dietary conditions studied. A fluorimetric assay with enzymic cycling was developed for the measurement of glucose 1-phosphate in HClO(4) extracts of rat liver. The calculated free cytoplasmic PP(i) concentration in nmol/g fresh wt. of liver was 2.3+/-0.3 in starved, 3.8+/-0.4 in fed, 4.9+/-0.6 in meal-fed and 5.2+/-0.4 in sucrose-re-fed animals. These values agree well with recently determined direct measurements of total PP(i) in rat liver and suggest that there is not a large amount of bound or metabolically inert PP(i) in rat liver. The cytoplasmic [ATP]/[AMP][PP(i)] ratio is 10(3) times the cytoplasmic [ATP]/[ADP][P(i)] ratio and varies differently with dietary state. The reaction PP(i)+H(2)O-->2P(i) catalysed by inorganic pyrophosphatase (EC 3.6.1.1) does not attain near-equilibrium in vivo. PP(i) should be considered as one of the group of small inorganic ions which is metabolically active and capable of exerting a controlling function in a number of important metabolic reactions.