Activity Of Rat Liver Phenylalanine Hydroxylase Research Articles

Phenylalanine-induced phosphorylation and activation of rat hepatic phenylalanine hydroxylase in vivo.

Rats were given intraperitoneal injections of 2 mCi of carrier-free 32Pi and substances known to activate liver phenylalanine hydroxylase. After 30 min, these animals were anesthetized and their livers removed for analysis of enzyme activity, 32Pi incorporation into immunoprecipitated phenylalanine hydroxylase and [gamma-32P]ATP specific activity. Following glucagon treatment, rat liver phenylalanine hydroxylase activity was stimulated more than 6-fold when assayed in the presence of the natural cofactor, tetrahydrobiopterin (BH4). Glucagon injection also resulted in an incorporation of 0.41 mol of 32Pi/mol of hydroxylase subunit (approximately 50,000 Da). In vivo stimulation of phenylalanine hydroxylase activity and 32Pi incorporation by glucagon had been previously observed in this laboratory (Donlon, J., and Kaufman, S. (1978) J. Biol. Chem. 253, 6657-6659). However, we show for the first time in the present study that in vivo treatment with phenylalanine alone results in a 4-fold increase in the BH4-dependent activity of phenylalanine hydroxylase concomitant with a significant incorporation of phosphate into phenylalanine hydroxylase (0.51 mol of 32Pi/mol of hydroxylase subunit). It is further demonstrated in vivo that the combined treatment with phenylalanine and glucagon results in a greater than 10-fold stimulation of BH4-dependent activity and the greatest level of 32Pi incorporation (0.75 mol of 32Pi/mol of hydroxylase subunit). Phenylalanine did not produce an elevation in plasma glucagon in these animals. A model is, thereby, proposed with respect to the ligand binding effects of phenylalanine on the state of phosphorylation and activation of phenylalanine hydroxylase. The significance of these regulatory roles are considered in light of the probable physiological environment of the enzyme.

Effect of glucagon on hepatic phenylalanine hydroxylase in vivo.

Moderate doses of glucagon (20 micrograms/kg I.V.) are sufficient to stimulate rat hepatic phenylalanine hydroxylase in vivo. In addition, the stimulation of the tetrahydrobiopterin-dependent phenylalanine hydroxylase activity in livers of animals fed on a high-protein diet has been correlated with an elevated phosphate content. The tetrahydrobiopterin-dependent hydroxylase activity in these animals can be further elevated by glucagon-stimulated phosphorylation. These results indicate that physiological changes in glucagon concentration modulate rat liver phenylalanine hydroxylase activity in vivo. The current understanding of the role of phosphorylation in regulating human phenylalanine hydroxylase is also considered.

Iron-dependent regulation of rat liver phenylalanine hydroxylase activity in vivo, in vitro, and in perfused liver.

Iron-dependent regulation of rat liver phenylalanine hydroxylase activity in vivo, in vitro, and in perfused liver.

In vivo inhibition of rat liver phenylalanine hydroxylase by p-chlorophenylalanine and esculin : Experimental model of phenylketonuria

The effect of p-chlorophenylalanine ( p-CPA) and esculin on the rat liver phenylalanine hydroxylase activity have been investigated. Results have demonstrated that both compounds are good inhibitors of the enzyme in vivo, the p-CPA being the strongest, and that esculin reinforces the inhibition produced by the former when both are administered together. l-Phenylalanine administration to rats treated with p-CPA plus esculin results in an enhancement of the levels of liver phenylalanine and its metabolites, which reach values similar to those found in tissues of phenylketonuric patients while tyrosine concentrations remain near the control values. All these results suggest that the use of the p-CPA plus esculin combination supplies an improved experimental model of phenylketonuria, which can be useful for biochemical studies.

Stimulation of rat liver phenylalanine hydroxylase activity by derivatives of vitamin E

The phenylalanine hydroxylase activity of rat liver preparations is enhanced by α-tocopheryl phosphate, α-tocopheryl succinate, and sodium dodecyl sulfate in vitro at 30°C and pH 7.4. The stimulation by α-tocopheryl phosphate (0.02 mM) is accompanied by a significant decrease in the Michaelis constant for phenylalanine and increases in the Michaelis constant for a synthetic pteridine cofactor and in the maximum velocity of the reaction. The possible relevance of these observations to phenylketonuria is discussed.

INFLUENCE OF AMINO ACIDS ON RAT LIVER PHENYLALANINE HYDROXYLASE ACTIVITY.

The influence of oral and of injected amino acids on rat liver phenylalanine hydroxylase activity has been studied in both short-term and extended experiments. Phenylalanine added to the diet in moderate amounts ordinarily causes an increase in enzyme activity, but when excess phenylalanine was included in the diet, it caused a marked decrease in the enzyme activity in rats fed ad libitum. Tryptophan or tyrosine added to the diet in the absence of added phenylalanine increased the enzyme activity in these rats. Excess tryptophan, but not tyrosine, could counteract the effect of phenylalanine on this enzyme after an 18-hr fast. There were dramatic differences in phenylalanine hydroxylase activity when various amino acids were injected. Injection of tryptophan or glycine decreased the phenylalanine hydroxylase activity in rat liver. Injections of other amino acids, notably phenylalanine and tyrosine, had no effect on the phenylalanine hydroxylase activity 5 hr after injection.