Phenylalanine hydroxylase: metabolic aspects.

Phenylalanine hydroxylase [L-phenylalanine,tetrahydropteridine :oxygen oxidoreductase (4-hydroxylating); EC 1.14.16.11 catalyses the first reaction in the irreversible catabolism of the essential amino acid phenylalanine. Studies of the isolated enzyme and of phenylalanine metabolism in viuo and in oitro support the view that the hydroxylase plays the major role in regulating phenylalanine disposal under many conditions. Phenylalanine and tyrosine (the immediate end product of hydroxylase activity) are both required for protein synthesis and probably share a single mechanism for entry into tissues. It is of some importance, therefore, that the balance between the concentrations of the two amino acids is maintained as physiological conditions change. The implication is that the activities of phenylalanine hydroxylase and of tyrosine aminotransferase (EC 2.6.1.9, the enzyme which initiates the oxidation of tyrosine, must be closely co-ordinated in vivo. In liver extracts, the activity of phenylalanine hydroxylase, as measured with optimal concentrations of a synthetic pteridine substrate and phenylalanine, is relatively high (23pmol/min per g wet wt. of tissue). Were such activity to be expressed in viuo, the body’s reserve of phenylalanine would be rapidly depleted. In practice, however, the rate with the natural pteridine, tetrahydrobiopterin, is less than 10% of this ‘maximum’ (Kaufman, 1971) and the low concentration of phenylalanine in the cell ensures that the rate is further decreased by an order of magnitude (Carr & Pogson, 1981). The concentrations of the two substrates are therefore important determinants of enzyme activity in vivo, although the precise role of the pteridine remains to be clarified (Milstien & Kaufman, 1975). In vitro, phenylalanine hydroxylase is phosphorylated by the cyclic AMP-dependent protein kinase (Abita et al., 1976; Donlon & Kaufman, 1980) and this phosphorylation has been shown in vivo (Donlon & Kaufman, 1978) and in liver cells (Abita et al., 1980) on exposure to glucagon. Changes in the activity of the enzyme expressed during incubation of cells with glucagon are only appreciable at low phenylalanine concentrations. Consistent with this, Shiman et al. (1982) suggest that phosphorylation does not activate the enzyme per se, but rather sensitizes it to the effect of phenylalanine acting as a stimulator at a site distinct from the catalytic site. It is apparent, however, that a phosphorylation-dephosphorylation cycle is potentially important in regulating the response of the enzyme to external stimuli, at least in the rat; the enzyme in human liver is reported not to undergo phosphorylation (Abita et al., 1983). Other relevant factors include, in the short term, the availability of iron to the liver cell (Shiman & Jefferson, 1982) and, over longer periods, changes in the enzyme’s total activity (Carr & Pogson, 1981 ; Donlon & Beirne, 1982) which parallel changes in hydroxylase protein concentration (M. A. Santana & C. I. Pogson, unpublished work). We have studied the relationship between phosphorylation of the enzyme and metabolic flux in simultaneous liver cell incubations. Guroff & Abramowitz (1967) devised a method for the