Biopterin cofactor biosynthesis: GTP cyclohydrolase, neopterin and biopterin in tissues and body fluids of mammalian species

Duch, David S.; Bowers, Seaton W.; Woolf, Jeffrey H.; Nichol, Charles A.

doi:10.1016/0024-3205(84)90541-1

Cited by 103 publications

(46 citation statements)

References 19 publications

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“…Their concluding hypothesis was that the responsiveness to BH 4 in patients with PAH deficiency was due to the suboptimal physiological concentrations of BH 4 normally present in hepatocytes and to the enhancement of the residual activity upon supplementation to higher than physiological concentrations. A hepatic concentration of BH 4 of 5-10 M has been measured (48)(49)(50). Michaelis-Menten kinetics tells us that the enzyme is not saturated at this concentration (K m ϭ 12-44 M; Table 2), and thus the activity will increase with increasing concentration of BH 4 .…”

Section: Discussionmentioning

confidence: 99%

Correction of kinetic and stability defects by tetrahydrobiopterin in phenylketonuria patients with certain phenylalanine hydroxylase mutations

Erlandsen

Pey

Gámez

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

152

162

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Phenylketonuria patients harboring a subset of phenylalanine hydroxylase (PAH) mutations have recently shown normalization of blood phenylalanine levels upon oral administration of the PAH cofactor tetrahydrobiopterin [(6R)-L-erythro-5,6,7,8-tetrahydrobiopterin (BH4)]. Several hypotheses have been put forward to explain BH 4 responsiveness, but the molecular basis for the corrective effect(s) of BH4 has not been understood. We have investigated the biochemical, kinetic, and structural changes associated with BH4-responsive mutations (F39L, I65T, R68S, H170D, E178G, V190A, R261Q, A300S, L308F, A313T, A373T, V388M, E390G, P407S, and Y414C). The biochemical and kinetic characterization of the 15 mutants studied points toward a multifactorial basis for the BH4 responsiveness; the mutants show residual activity (>30% of WT) and display various kinetic defects, including increased Km (BH4) and reduced cooperativity of substrate binding, but no decoupling of cofactor (BH4) oxidation. For some, BH4 seems to function through stabilization and protection of the enzyme from inactivation and proteolytic degradation. In the crystal structures of a phenylketonuria mutant, A313T, minor changes were seen when compared with the WT PAH structures, consistent with the mild effects the mutant has upon activity of the enzyme both in vitro and in vivo. Truncations made in the A313T mutant PAH form revealed that the N and C termini of the enzyme influence active site binding. Of fundamental importance is the observation that BH4 appears to increase Phe catabolism if at least one of the two heterozygous mutations has any residual activity remaining.

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Section: Discussionmentioning

confidence: 99%

Correction of kinetic and stability defects by tetrahydrobiopterin in phenylketonuria patients with certain phenylalanine hydroxylase mutations

Erlandsen

Pey

Gámez

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

152

162

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“…In humans, cytokine-induced stimulation of GTP cyclohydrolase I can be easily detected in vivo by measur- triphosphate accumulates in human but not in murine cells due to the comparatively low 6-pyruvoyl tetrahydropterin synthase activity found in human cells [4,6], thus explaining excretion of neopterin in humans but not in rodents [7]. Among human cells, the ratio of GTP ~yclohydrolase I to 6-pyruvoyl tetrahydropterin synthase after cytokine treatment is highest in macrophages which leads to production of an about 50-fold excess of neopterin over biopterin [6,8].…”

Section: Introductionmentioning

confidence: 99%

Induction of GTP cyclohydrolase I by bacterial lipopolysaccharide in the rat

et al. 1993

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A 2-to 3-fold increase of GTP cyclohydrolase I (E.C. 3.5.4.16). the key enzyme of tetrahydrobiopterin biosynthesis from GTP, was observed in cerebellum, remaining brain, liver, spleen, and adrenal gland of rats treated with a single dose of lipopolysaccha~de (LPS). This led to increased biopte~n levels in tissues but not in plasma. Parallel induction of nitric oxide (NO) synthase was indicated by a IO-to I~-fold increase of plasma nitrate levels 6 and 12 hours after injection of LPS. Furthe~ore, systolic blood pressure was reduced significantly by 23%. Our results demonstrate induction of tetrahydrobiopterin biosynthesis after LPS treatment in vivo.

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“…For cruder material and in some experiments with purified enzyme, activity was determined by measurement of neopterin produced after oxidation and dephosphorylation of the enzymatic product, dihydroneopterin triphosphate, by modifications of previously described methods (19). In brief, samples to be assayed were buffer-exchanged into buffer A (50 mM KPO 4 (pH 7.4), 1 mM EDTA, 1 mM DTT, 0.1% Tween 20, 10% glycerol) by the spin column method through Sephadex G25-M, and aliquots were incubated at 37°C for various times (usually 15 min) in a total volume of 50 l containing 0.5 mM Tris-HCl (pH 7.4), 1 mM DTT, 50 g of BSA, and 1 mM GTP.…”

Section: Enzyme Assaysmentioning

confidence: 99%

Purification and Cloning of the GTP Cyclohydrolase I Feedback Regulatory Protein, GFRP

Milstien

Jaffe

Kowlessur

et al. 1996

Journal of Biological Chemistry

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The activity of GTP cyclohydrolase I, the initial enzyme of the de novo pathway for biosynthesis of tetrahydrobiopterin, the cofactor required for aromatic amino acid hydroxylations and nitric oxide synthesis, is sensitive to end-product feedback inhibition by tetrahydrobiopterin. This inhibition by tetrahydrobiopterin is mediated by the GTP cyclohydrolase I feedback regulatory protein GFRP, previously named p35 (Harada, T., Kagamiyama, H., and Hatakeyama, K. (1993) Science 260, 1507-1510), and L-phenylalanine specifically reverses the tetrahydrobiopterin-dependent inhibition. As a first step in the investigation of the physiological role of this unique mechanism of regulation, a convenient procedure has been developed to co-purify to homogeneity both GTP cyclohydrolase I and GFRP from rat liver. GTP cyclohydrolase I and GFRP exist in a complex which can be bound to a GTP-affinity column from which GTP cyclohydrolase I and GFRP are separately and selectively eluted. GFRP is dissociated from the GTP agarose-bound complex with 0.2 M NaCl, a concentration of salt which also effectively blocks the tetrahydrobiopterin-dependent inhibitory activity of GFRP. GTP cyclohydrolase I is then eluted from the GTP-agarose column with GTP. Both GFRP and GTP cyclohydrolase I were then purified separately to near homogeneity by sequential high performance anion exchange and gel filtration chromatography. GFRP was found to have a native molecular mass of 20 kDa and consist of a homodimer of 9.5-kDa subunits. Based on peptide sequences obtained from purified GFRP, oligonucleotides were synthesized and used to clone a cDNA from a rat liver cDNA library by polymerase chain reaction-based methods. The cDNA contained an open reading frame that encoded a novel protein of 84 amino acids (calculated molecular mass 9665 daltons). This protein when expressed in Escherichia coli as a thioredoxin fusion protein had tetrahydrobiopterin-dependent GTP cyclohydrolase I inhibitory activity. Northern blot analysis indicated the presence of an 0.8-kilobase GFRP mRNA in most rat tissues, the amounts generally correlating with levels of GTP cyclohydrolase I and tetrahydrobiopterin. Thus, mRNA levels were relatively high in liver and kidney and somewhat lower in testis, heart, brain, and lung. These results suggest that GFRP is widely expressed and may play a role in regulating not only phenylalanine metabolism in the liver, but also the production of biogenic amine neurotransmitters as well as nitric oxide synthesis.

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Biopterin cofactor biosynthesis: GTP cyclohydrolase, neopterin and biopterin in tissues and body fluids of mammalian species

Cited by 103 publications

References 19 publications

Correction of kinetic and stability defects by tetrahydrobiopterin in phenylketonuria patients with certain phenylalanine hydroxylase mutations

Correction of kinetic and stability defects by tetrahydrobiopterin in phenylketonuria patients with certain phenylalanine hydroxylase mutations

Induction of GTP cyclohydrolase I by bacterial lipopolysaccharide in the rat

Purification and Cloning of the GTP Cyclohydrolase I Feedback Regulatory Protein, GFRP

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