p-Chlorphenylalanine effect on phenylalanine hydroxylase in hepatoma cells in culture.
We have investigated the p-chlorophenylalanine-dependent loss of phenylalanine hydroxylase activity in cultured hepatoma cells. The similarity of the effect of p-chlorophenylalanine on phenylalanine hydroxylase in the hepatoma cells and that reported from studies in vivo indicates that the loss of phenylalanine hydroxylase activity is due to a direct interaction of the amino acid analogue with the liver. We can find no evidence that the loss of phenylalanine hydroxylase activity is due to: a direct inactivation of the hydroxylase by p-chlorophenylalanine or an inhibitor produced by p-chlorophenylalanine treatment; an effect similar to that of p-fluorophenylalanine; or leakage of enzyme from the cells during p-chlorophenylalanine treatment. The data presented indicate: (a) the p-chlorophenylalanine effect is rather specific for phenylalanine hydroxylase; (b) following p-chlorophenylalanine removal, new protein synthesis is necessary for restoration of the hydroxylase activity; (c) the rate of loss of phenylalanine hydroxylase activity after the addition of p-chlorophenylalanine is much faster than the rate of restoration of the hydroxylase activity after removal of p-chlorophenylalanine; (d) even in the presence of p-chlorophenylalanine, hydrocortisone greatly stimulates the hydroxylase activity; (e) the cell density-dependent increase of phenylalanine hydroxylase activity is blocked by p-chlorophenylalanine. A discussion of the possible mechanisms of p-chlorophenylalanine-dependent loss of phenylalanine hydroxylase is presented. To measure very low leanine-dependent loss of phenylalanine hydroxylase is presented. To measure very low levels of phenylalanine hydroxylase activity, a new procedure, based on isotope dilution, was developed for isolating the tyrosine formed during the enzymatic reaction.
D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000201:enzymology*; D002478:Cells, Cultured; D010134:Fenclonine / Q000494:pharmacology*; D006854:Hydrocortisone / Q000494:pharmacology; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000201:enzymology; D008113:Liver Neoplasms; D008244:Lysophosphatidylcholines / Q000494:pharmacology; D009374:Neoplasms, Experimental / Q000201:enzymology; D010651:Phenylalanine Hydroxylase / Q000378:metabolism*; D011622:Pterins / Q000494:pharmacology; D051381:Rats; D013997:Time Factors; D014444:Tyrosine Transaminase / Q000378:metabolism
D008244:Lysophosphatidylcholines; D011622:Pterins; D007770:L-Lactate Dehydrogenase; D010651:Phenylalanine Hydroxylase; D014444:Tyrosine Transaminase; D010134:Fenclonine; D006854:Hydrocortisone