The effect of phenylalanine on DOPA synthesis in PC12 cells.
DOPA synthesis from phenylalanine was studied in PC12 cells incubated with m-hydroxybenzylhydrazine, to inhibit aromatic L-amino acid decarboxylase. DOPA synthesis rose with increasing concentrations of either phenylalanine or tyrosine; maximal rates (approximately 55 pmol/min/mg protein for tyrosine; approximately 40 pmol/min/mg protein for phenylalanine) occurred at a medium concentration of approximately 10 microM for either amino acid. The Km for either amino acid was about 1 microM (medium concentration). At tyrosine concentrations above 30 microM, DOPA synthesis declined; inhibition was observed at higher concentrations for phenylalanine (> or =300 microM). These effects were most notable in the presence of 56 mM potassium. Measurements of intracellular phenylalanine and tyrosine suggested the Km for either amino acid is 20-30 microM; maximal synthesis occurred at 120-140 microM. In the presence of both phenylalanine and tyrosine, DOPA synthesis was inhibited by phenylalanine only at a high medium concentration (1000 microM), regardless of medium tyrosine concentration. The inhibition of DOPA synthesis by high medium tyrosine concentrations was antagonized by high medium phenylalanine concentrations (100, 1000 microM). Together, the findings indicate that for PC12 cells, phenylalanine can be a significant substrate for tyrosine hydroxylase, is a relatively weak inhibitor of the enzyme, and at high concentrations can antagonize substrate inhibition by tyrosine.
Department of Molecular Biology and Biochemistry, University of Pittsburgh School of Medicine, PA, USA.
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DePietro
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Fernstrom
D000818:Animals; D004295:Dihydroxyphenylalanine / Q000096:biosynthesis*; D006900:Hydroxylation; D016716:PC12 Cells; D010649:Phenylalanine / Q000494:pharmacology*; D051381:Rats; D015203:Reproducibility of Results; D014443:Tyrosine / Q000378:metabolism
D014443:Tyrosine; D010649:Phenylalanine; D004295:Dihydroxyphenylalanine