tetrahydropterin and sapropterin

This work had two purposes: (i) to determine in vivo whether liver phenylalanine hydroxylase (PAH) is regulated by its substrates phenylalanine and tetrahydrobiopterin (BH4) as studies with purified enzyme suggest and (ii) to investigate in vivo the relationship between PAH activity and BH4 turnover. We found there are two BH4 pools in hepatocytes, one that is metabolically available (free BH4) and one that is not (bound BH4). Bound BH4 appears bound to PAH; the PAH-BH4 complex has much 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 concentration is determined by the state of activation and activity of PAH. In hepatocytes, BH4 and PAH (subunit) concentrations are equal, all intracellular BH4 appears to be available to PAH, and free BH4 turns over rapidly (t1/2 approximately 1 hr). There is no evidence for feedback inhibition of BH4 synthesis; the BH4 synthetic rate appears high when free BH4 concentration is high and low when free BH4 is low. The data provide support in vivo that phenylalanine and BH4 are positive and negative regulators of the activity and activation state of PAH in the proposed manner; they also imply that regulation of BH4 turnover and PAH activity are linked processes, which are both controlled by phenylalanine concentration.

Topics: Animals; Binding, Competitive; Biopterins; Cells, Cultured; Enzyme Activation; Kinetics; Liver; Phenylalanine; Phenylalanine Hydroxylase; Pterins; Rats; Water

The conversion of dihydroneopterin triphosphate in the presence of 6-pyruvoyl tetrahydropterin synthase was followed by 1H-NMR spectroscopy. The interpretation of the spectra of the product is unequivocal: they show formation of a tetrahydropterin system carrying a stereospecifically oriented substituent at the asymmetric C(6) atom. The spectra are compatible with formation of a (3')-CH3 function, and with complete removal of the 1' and 2' hydrogens of dihydroneopterin triphosphate. The fast-atom-bombardment/mass spectrometry study of the same product yields a [M + H]+ ion at m/z 238 compatible with the structure of 6-pyruvoyl tetrahydropterin. The data support the proposed structure of 6-pyruvoyl tetrahydropterin as a key intermediate in the biosynthesis of tetrahydrobiopterin.

Topics: Alcohol Oxidoreductases; Biopterins; Magnetic Resonance Spectroscopy; Mass Spectrometry; Models, Chemical; Neopterin; Oxidation-Reduction; Phosphorus-Oxygen Lyases; Pteridines; Pterins; Stereoisomerism

Using reversed-phase high-performance liquid chromatography with electrochemical detection we have demonstrated the occurrence of 5,6,7,8-tetrahydropterin and 5,6,7,8-tetrahydrobiopterin in Drosophila melanogaster. The former is the first time that has been detected in vivo. The identification has been based on the retention times, hydrodinamic voltagrams and the differential concentration in three strains of Drosophila melanogaster. Compared to the wild type, the Punch2 mutant has diminished levels of both pteridines, whereas Henna-recessive3 lacks completely tetrahydropterin and has increased levels of tetrahydrobiopterin, as expected according to their biochemical lesions.

Topics: Animals; Biopterins; Chromatography, High Pressure Liquid; Drosophila melanogaster; Kidney; Pterins; Rats

It is known that the first step in the de novo synthesis of tetrahydrobiopterin from GTP is the conversion of GTP to dihydroneopterin triphosphate. Recent evidence supports the conclusion that beyond this first step, the pterin intermediates in the pathway are all at the tetrahydro level of reduction. We have now shown that partially purified fractions from rat liver, rat brain and bovine adrenal medulla catalyze the conversion of dihydroneopterin triphosphate to tetrahydrobiopterin, as well as to the putative intermediates in the pathway, 6-pyruvoyl-tetrahydropterin and 6-lactoyl-tetrahydropterin. Results of both enzymatic and chemical studies support the assigned structures for the latter two tetrahydropterins. We have also purified extensively from brain an enzyme, distinct from sepiapterin reductase, that catalyzes the TPNH-dependent reduction of 6-pyruvoyl-tetrahydropterin to 6-lactoyl-tetrahydropterin. The role of this reductase in tetrahydrobiopterin synthesis has not yet been established.

Topics: Adrenal Medulla; Animals; Biopterins; Brain; Cattle; Chemical Phenomena; Chemistry; In Vitro Techniques; Kinetics; Liver; Neopterin; Pteridines; Pterins; Rats

Isolation of two kinetically competent tetrahydropterin intermediates on the de novo biosynthetic pathway to tetrahydrobiopterin is reported. The compounds were detected in HPLC chromatograms by electrochemical oxidation at 270 mv. The hydrodynamic voltamograms of the compounds are indistinguishable from those of tetrahydropterins . Both require Mg+ + for biosynthesis from dihydroneopterin triphosphate. One compound is produced in the absence of NADPH; the other requires NADPH for synthesis. Enzymatic conversion of the former into the latter intermediate requires NADPH. Conversion of either intermediate into tetrahydrobiopterin also requires NADPH. Mg+ +, which is required for the biosynthesis of the intermediates, is not needed for their conversion to tetrahydrobiopterin. Neither compound coelutes with 6-lactoyl tetrahydropterin, the tetrahydropterin analog of sepiapterin.

Topics: Adrenal Medulla; Animals; Biopterins; Cattle; Chromatography, High Pressure Liquid; Electrochemistry; Kinetics; NADP; Oxidation-Reduction; Pteridines; Pterins

Substantial amounts of tetrahydrobiopterin and 6-methyltetrahydropterin can be detected in CSF when these pterins are given peripherally to patients with hyperphenylalaninemia due to defective biopterin synthesis. Results of this study suggest that administration of either of these pterins in proper doses may prove to be a treatment not only for the impaired peripheral phenylalanine metabolism, but also for the neurologic disorders that are characteristic of the variant forms of hyperphenylalaninemia due to defective tetrahydrobiopterin synthesis or metabolism.

Topics: Biopterins; Brain; Child; Child, Preschool; Female; Humans; Male; Neopterin; Phenylalanine; Phenylketonurias; Pteridines; Pterins