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. 1984 Apr 1;219(1):79–85. doi: 10.1042/bj2190079

Phenylalanine hydroxylase in liver cells. Correlation of glucagon-stimulated enzyme phosphorylation with expressed activity.

M J Fisher, C I Pogson
PMCID: PMC1153450  PMID: 6721866

Abstract

Phenylalanine is transported rapidly into, but is not concentrated by, liver cells. Glucagon increased flux through phenylalanine hydroxylase; a half-maximal response was obtained at 0.7 nM. Under control conditions, 0.2-0.3 mol of phosphate were incorporated per mol of subunit of the hydroxylase at steady state. Glucagon increased this incorporation of phosphate into the hydroxylase to a maximal value of approx. 0.6 mol of phosphate per subunit; a half-maximal response was obtained at 0.3 nM. Glucagon, added simultaneously with [32P]Pi to liver cells, inhibited incorporation of 32P into the enzyme. The effects of glucagon were reproduced with dibutyryl cyclic AMP. Changes in phosphorylation correlated closely with changes in flux through phenylalanine hydroxylase in cell incubations.

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Selected References

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  1. Abita J. P., Chamras H., Rosselin G., Rey F. Hormonal control of phenylalanine hydroxylase activity in isolated rat hepatocytes. Biochem Biophys Res Commun. 1980 Feb 12;92(3):912–918. doi: 10.1016/0006-291x(80)90789-5. [DOI] [PubMed] [Google Scholar]
  2. Abita J. P., Milstien S., Chang N., Kaufman S. In vitro activation of rat liver phenylalanine hydroxylase by phosphorylation. J Biol Chem. 1976 Sep 10;251(17):5310–5314. [PubMed] [Google Scholar]
  3. Baur H., Heldt H. W. Transport of hexoses across the liver-cell membrane. Eur J Biochem. 1977 Apr 1;74(2):397–403. doi: 10.1111/j.1432-1033.1977.tb11404.x. [DOI] [PubMed] [Google Scholar]
  4. Blair J. B., Cimbala M. A., James M. E. Hepatic pyruvate kinase. Quantitative measurements of phosphorylation in vitro and in the isolated rat hepatocyte. J Biol Chem. 1982 Jul 10;257(13):7595–7602. [PubMed] [Google Scholar]
  5. Carr F. P., Pogson C. I. Phenylalanine metabolism in isolated rat liver cells. Effects of glucagon and diabetes. Biochem J. 1981 Sep 15;198(3):655–660. doi: 10.1042/bj1980655. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chen R. F. Removal of fatty acids from serum albumin by charcoal treatment. J Biol Chem. 1967 Jan 25;242(2):173–181. [PubMed] [Google Scholar]
  7. Christensen H. N. Some special kinetic problems of transport. Adv Enzymol Relat Areas Mol Biol. 1969;32:1–20. doi: 10.1002/9780470122778.ch1. [DOI] [PubMed] [Google Scholar]
  8. Dickson A. J., Marston F. A., Pogson C. I. Tyrosine aminotransferase as the rate-limiting step for tyrosine catabolism in isolated rat liver cells. FEBS Lett. 1981 May 5;127(1):28–32. doi: 10.1016/0014-5793(81)80333-x. [DOI] [PubMed] [Google Scholar]
  9. Dickson A. J., Pogson C. I. The metabolic integrity of hepatocytes in sustained incubations. FEBS Lett. 1977 Nov 1;83(1):27–32. doi: 10.1016/0014-5793(77)80634-0. [DOI] [PubMed] [Google Scholar]
  10. Donlon J., Kaufman S. Glucagon stimulation of rat hepatic phenylalanine hydroxylase through phosphorylation in vivo. J Biol Chem. 1978 Oct 10;253(19):6657–6659. [PubMed] [Google Scholar]
  11. Donlon J., Kaufman S. Relationship between the multiple forms of rat hepatic phenylalanine hydroxylase and degree of phosphorylation. J Biol Chem. 1980 Mar 10;255(5):2146–2152. [PubMed] [Google Scholar]
  12. Edmondson J. W., Lumeng L., Li T. K. Comparative studies of alanine and alpha-aminoisobutyric acid uptake by freshly isolated rat liver cells. J Biol Chem. 1979 Mar 10;254(5):1653–1658. [PubMed] [Google Scholar]
  13. Garrison J. C., Wagner J. D. Glucagon and the Ca2+-linked hormones angiotensin II, norepinephrine, and vasopressin stimulate the phosphorylation of distinct substrates in intact hepatocytes. J Biol Chem. 1982 Nov 10;257(21):13135–13143. [PubMed] [Google Scholar]
  14. Guroff G., Abramowitz A. A simple radioisotope assay for phenylalanine hydroxylase. Anal Biochem. 1967 Jun;19(3):548–555. doi: 10.1016/0003-2697(67)90245-x. [DOI] [PubMed] [Google Scholar]
  15. Guroff G., Daly J. W., Jerina D. M., Renson J., Witkop B., Udenfriend S. Hydroxylation-induced migration: the NIH shift. Recent experiments reveal an unexpected and general result of enzymatic hydroxylation of aromatic compounds. Science. 1967 Sep 29;157(3796):1524–1530. doi: 10.1126/science.157.3796.1524. [DOI] [PubMed] [Google Scholar]
  16. Hasegawa H., Parniak M., Kaufman S. Determination of the phosphate content of purified proteins. Anal Biochem. 1982 Mar 1;120(2):360–364. doi: 10.1016/0003-2697(82)90358-x. [DOI] [PubMed] [Google Scholar]
  17. Hawkins P. T., Michell R. H., Kirk C. J. A simple assay method for determination of the specific radioactivity of the gamma-phosphate group of 32P-labelled ATP. Biochem J. 1983 Mar 15;210(3):717–720. doi: 10.1042/bj2100717. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  19. Le Cam A., Freychet P. Neutral amino acid transport. Characterization of the A and L systems in isolated rat hepatocytes. J Biol Chem. 1977 Jan 10;252(1):148–156. [PubMed] [Google Scholar]
  20. Marston F. A., Pogson C. I. A simple and rapid assay for tyrosine aminotransferase. FEBS Lett. 1977 Nov 15;83(2):277–280. doi: 10.1016/0014-5793(77)81022-3. [DOI] [PubMed] [Google Scholar]
  21. McGivan J. D., Bradford N. M., Mendes-Mourão J. The transport of branched-chain amino acids into isolated rat liver cells. FEBS Lett. 1977 Aug 15;80(2):380–384. doi: 10.1016/0014-5793(77)80481-x. [DOI] [PubMed] [Google Scholar]
  22. Milstien S., Abita J. P., Chang N., Kaufman S. Hepatic phenylalanine 4-monooxygenase is a phosphoprotein. Proc Natl Acad Sci U S A. 1976 May;73(5):1591–1593. doi: 10.1073/pnas.73.5.1591. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. OXENDER D. L., CHRISTENSEN H. N. DISTINCT MEDIATING SYSTEMS FOR THE TRANSPORT OF NEUTRAL AMINO ACIDS BY THE EHRLICH CELL. J Biol Chem. 1963 Nov;238:3686–3699. [PubMed] [Google Scholar]
  24. Parniak M., Hasegawa H., Wilgus H., Kaufman S. On the phosphate content of rat liver phenylalanine hydroxylase purified by hydrophobic chromatography. Biochem Biophys Res Commun. 1981 Mar 31;99(2):707–714. doi: 10.1016/0006-291x(81)91801-5. [DOI] [PubMed] [Google Scholar]
  25. Phillips R. S., Iwaki M., Kaufman S. Ligand effects on the limited proteolysis of phenylalanine hydroxylase: evidence for multiple conformational states. Biochem Biophys Res Commun. 1983 Feb 10;110(3):919–925. doi: 10.1016/0006-291x(83)91050-1. [DOI] [PubMed] [Google Scholar]
  26. Seglen P. O., Gordon P. B. Effects of lysosomotropic monoamines, diamines, amino alcohols, and other amino compounds on protein degradation and protein synthesis in isolated rat hepatocytes. Mol Pharmacol. 1980 Nov;18(3):468–475. [PubMed] [Google Scholar]
  27. Shiman R., Gray D. W., Pater A. A simple purification of phenylalanine hydroxylase by substrate-induced hydrophobic chromatography. J Biol Chem. 1979 Nov 25;254(22):11300–11306. [PubMed] [Google Scholar]
  28. Shiman R., Jefferson L. S. Iron-dependent regulation of rat liver phenylalanine hydroxylase activity in vivo, in vitro, and in perfused liver. J Biol Chem. 1982 Jan 25;257(2):839–844. [PubMed] [Google Scholar]
  29. Shiman R., Mortimore G. E., Schworer C. M., Gray D. W. Regulation of phenylalanine hydroxylase activity by phenylalanine in vivo, in vitro, and in perfused rat liver. J Biol Chem. 1982 Oct 10;257(19):11213–11216. [PubMed] [Google Scholar]
  30. Shiman R. Relationship between the substrate activation site and catalytic site of phenylalanine hydroxylase. J Biol Chem. 1980 Nov 10;255(21):10029–10032. [PubMed] [Google Scholar]
  31. Smith S. A., Pogson C. I. The metabolism of L-tryptophan by isolated rat liver cells. Effect of albumin binding and amino acid competition on oxidatin of tryptophan by tryptophan 2,3-dioxygenase. Biochem J. 1980 Mar 15;186(3):977–986. doi: 10.1042/bj1860977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. WAALKES T. P., UDENFRIEND S. A fluorometric method for the estimation of tyrosine in plasma and tissues. J Lab Clin Med. 1957 Nov;50(5):733–736. [PubMed] [Google Scholar]
  33. Wicks W. D., Su J. L. Relationship between phosphorylation of tyrosine aminotransferase and regulation of its synthesis by cyclic AMP and hormones. J Cyclic Nucleotide Res. 1978 Apr;4(2):113–122. [PubMed] [Google Scholar]

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