COORDINATE REGULATION OF TETRAHYDROBIOPTERIN TURNOVER AND PHENYLALANINE-HYDROXYLASE ACTIVITY IN RAT-LIVER CELLS

This work had two purposes: (i) to determine in vivo whether liver phe nylalanine hydroxylase (PAH) is regulated by its substrates phenylalan ine and tetrahydrobiopterin (BH4) as studies with purified enzyme sugg est and (ii) to investigate in vivo the relationship between PAH activ ity and BH4 turnover. We found there are two BH4 pools in hepatocytes, one that is metabolically available (free BH4) and one that is not (b ound BH4). Bound BH4 appears bound to PAH; the PAH-BH4 complex has muc h less catalytic activity and is less readily phenylalanine activated than uncomplexed enzyme. Interconversion of activated and unactivated PAH and bound and free BH4 is driven by phenylalanine; and free BH4 co ncentration is determined by the state of activation and activity of P AH. In hepatocytes, BH4 and PAH (subunit) concentrations are equal, al l intracellular BH4 appears to be available to PAH, and free BH4 turns over rapidly (t(1/2) approximate to 1 hr). There is no evidence for f eedback inhibition of BH4 synthesis; the BH4 synthetic rate appears hi gh when free BH4 concentration is high and low when free BH4 is low. T he data provide support in vivo that phenylalanine and BH4 are positiv e and negative regulators of the activity and activation state of PAH in the proposed manner; they also imply that regulation of BH4 turnove r and PAH activity are linked processes, which are both controlled by phenylalanine concentration.