Sepiapterin reductase

Fig. 6.1.8a-c HPLC of the yellow-fluorescing pterins, a Standard mixture b control cerebrospinal fluid (CSF) c CSF sepiapterin reductase deficiency. HS -Hydroxyse-piapterin, S sepiapterin, X xanthopterin... 

Bonafe L, Thony B, Penzien JM, Czarnecki B, Blau N (2001) Mutations in the sepiapterin reductase gene cause a novel tetrahydrobiopterin-dependent monoamine neurotransmitter deficiency without hyperphenylalaninemia. Am J Hum Genet 69 269-277... 

Sueoka T, Katoh S (1982) Purification and characterization of sepiapterin reductase from rat erythrocytes. Biochim Biophys Acta 717 265-271... 

Citron BA, Milstien S, Gutierrez JC, Levine RA, Yanak BL, Kaufman S (1990) Isolation and expression of rat liver sepiapterin reductase cDNA. Proc Natl Acad Sci U S A 87 6436-6440... 

Ichinose H, Katoh S, Sueoka T, Titani K, Fujita K, Nagatsu T (1991) Cloning and sequencing of cDNA encoding human sepiapterin reductase - an enzyme involved in tetrahydrobiop-terin biosynthesis. Biochem Biophys Res Commun 179 183-189... 

Ferre J, Naylor EW (1988) Sepiapterin reductase in human amniotic and skin fibroblasts, chorionic villi, and various blood fractions. Clin Chim Acta 174 271-282... 

Blau N, Bonafe L, Thony (2001) Tetrahydrobiopterin deficiencies without hyperphenyl-alaninemia diagnosis and genetics of Dopa-responsive dystonia and sepiapterin reductase deficiency. Mol Genet Metab 74 172-185... 

Enzymatic reactions in the side chains of pteridines are recommended for such cases where the chemical transformations are difficult and nonselective. A convincing example of this approach is the enzymatic isomerization of sepiapterin (6-lactoyl-7,8-dihydropterin) (297) into 6-(l-hydroxy-2-oxopropyl)-7,8-dihydropterin (298) which had been believed to be too labile to actually exist (Equation (13)) <89MI 718-06). This product, which was formed by the isomerase activity of sepiapterin reductase, was detectable electrochemically at 500 mV and fluorometrically on HPLC analysis. 

Sepiapterin reductase is included in the assay to allow the product to proceed to tetrahydrobiopterin, which is then oxidized to biopterin, which is detected by fluorescence. 

The standard reaction mixture was composed of 5 /aL of Tris-HCl (pH 7.4), 5 /aL of 40 mM NADPH, 5 /aL of sepiapterin reductase (activity of 400 nmol/min/mL), and 65 /aL of cell extract (10-200 /Ag of protein). The reaction was started by the addition of 20 /aL of 0.4 mM 7,8-dihydroneopterin triphosphate. After 30 to 90 minutes of incubation in the dark at 37°C, the reaction was terminated by the addition of 50 /aL of a mixture of 0.2 M HC1 and 0.02 M KI-I2 (11, v/v). The resulting mixture was incubated for 1 hour in the dark to allow oxidation of tetrahydrobiopterin to biopterin. Excess iodine was destroyed by the addition of 50 /aL of 0.02 M ascorbic add. An aliquot of the mixture was applied to a solid phase cartridge (SCX from Analytichem) that had been preequilibrated with 0.1 M H3PO4. The sample was forced through the cartridge with air pressure. The cartridge was then washed with 0.5 mL of 0.1 M H3PO4. The eluates were used for HPLC analysis. Assays were linear with up to 150 fig of cellular protein and 90 minutes of incubation. 

Figure 10.2. Biosynthesisoffolicacidandtetrahydrobiopterin.GTPcyclohydrolasel.EC 3.5.4.16 dihydropteroate synthase, EC 2.5.1.15 pyruvoyl-tetrahydrobiopterin synthase, EC 4.6.1.10 and sepiapterin reductase, EC 1.1.1.153.

The formation of biopterin involves dephosphorylation and reduction of the side chain of dihydroneopterin triphosphate, followed by inversion of the conformation of the two hydroxyl groups, by way of intermediate oxidation to (symmetrical) oxo-groups, catalyzed by sepiapterin reductase. 

These oxidation reactions require oxygen (O2) and tetrahydrobiopterin as a cofactor. Thus, as shown in Scheme 13.39, 7,8-dihydroneopterin 3 -triphosphate (generated from guanosine triphosphate [GTP] as seen in Scheme 12.118) is converted to 6-pyruvoyl-5,6,7,8-tetrahydropterin by an elimination reaction and two keto-enol isomerizations. The process is catalyzed by the enzyme 6-pyruvoyltetra-hydropterin synthase (EC 4.2.3.12). Then, via an intermediate, written as an equilibrium between a-hydroxyketones (named dihydrosepiapterin) linked by a common enol, reduction to tetrahydrobiopterin is effected (in two separate steps) by 2 equivalents of NADPH used by the enzyme sepiapterin reductase (EC 1.1.1.153). Tetrahydrobiopterin is the cofactor involved in the National Institutes of Health (NIH) shift (cf. Chapter 6) pathway used by the iron-containing enzyme phenylalanine 4-monooxygenase (EC 1.14.16.1) to convert phenylalanine (Phe, F) to tyrosine (Tyr, Y) and is converted to (6i )-6-(L-erythro-l,2-dihydroxypropyl)-5,6,7,8-tetrahydro-4a-hydroxypterin in the process. 

A-ACPEA and A-ACTYR were compared for their inhibitory effects on sepiapterin reductase, a catalyzing enzyme for synthesis of tetrahydrobiopterin which is a required cofactor for phenylalanine, tyrosine, and tryptophan hydroxylases. hi MOLT-4 T cell leukemia and MCF-7 breast cancer cell lines, both A-ACPEA and A-ACTYR (Af-acetyl-p-tyramine) showed poor inhibitory effects on sepiapterin reductase, whereas Af-acetyl-m-tyramine was a strong inhibitor [42]. 

Smith GBC, Duch DS, Edelstein MP, Bigham EC (1992) New inhibitors of sepiapterin reductase. J Biol Chem 267(8) 5599-5607... 

Senecio spp., 1384, 1553 Seneciphylline, 1061, 1062, 1178, 1386 Senegin III, 3217 Senile plaques, 3712, 3716 Sensitivity enhancement, 1105 Sensory analysis, 2267 Sensory characteristic, 2906 Sensory perception, 2264 Sepiapterin reductase, 1209 Sepsis, 1306 S-Eq producers, 2402 Sequence tags, 2954 S-equol (5-Eq), 2398 SERCA, 3077, 3090, 3091, 3093, 3095 Sergeolide, 3778 L-Serine, 942 Serine protease, 987, 988 Serjania meridionalis, 4094 Serotonergic (5-HT), 3709, 3711 neurons, 1516 system, 1309 Serotonin, 598, 686, 4068 Serpentine, 1126 Sertiiumer, 1368,1373 Sesame oil, 4065 Sesamin, 4065... 

IWo disorders of BH4 metabolism may present without hyperphenylalaninemia. These are Dopa-responsive dystonia (DRD Segawa disease) and sepiapterin reductase (SR) deficiency. While DRD is caused by a mutation in the GTPCH gene and is inherited in an autosomal dominant manner, SR deficiency is an autosomal recessive trait. Both diseases evidence severe biogenic amines deficiencies. DRD usually presents with a dystonic gait and diurnal variation. At least two reports describe heteroallelic patients with DRD suggesting a wide spectrum of GTPCH variants. 

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