Publications

• Purification and characterization of sepiapterin reductase from rat erythrocytes.

  Sueoka T., Katoh S.

  Sepiapterin reductase from rat erythrocyte hemolysate was purified 2000-fold to apparent homogeneity with 30% yield. The specific activity of the purified enzyme was 18 units/mg protein, and its molecular weight was 55 000. The enzyme consists of two identical subunits, each of which has a molecul ... >> More

  Sepiapterin reductase from rat erythrocyte hemolysate was purified 2000-fold to apparent homogeneity with 30% yield. The specific activity of the purified enzyme was 18 units/mg protein, and its molecular weight was 55 000. The enzyme consists of two identical subunits, each of which has a molecular weight of 27 500. The enzyme showed a single peak by isoelectric focusing with a pI of 4.9 and partial specific volume of 0.73 cm3/g. The amino acid composition was determined. pH optimum of the enzyme was 5.5. The equilibrium constant of 2.2.10(9) of the enzyme showed that the equilibrium lies much in favor of dihydrobiopterin formation from sepiapterin in rat erythrocytes. From steady-state kinetic measurements, ordered bi-bi mechanism was proposed to the reaction of sepiapterin reductase in which NADPH binds to free enzyme and sepiapterin binds next. NADP+ is released after the release of dihydrobiopterin. The Km values for sepiapterin and NADPH were 15.4 microM and 1.7 microM, respectively, and the Vmax value was 21.7 mumol/min per mg. << Less

  Biochim Biophys Acta 717:265-271(1982) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Expression, purification, crystallization and preliminary X-ray analysis of sepiapterin reductase from Chlorobium tepidum.

  Supangat S., Choi Y.K., Park Y.S., Son D., Han C.D., Lee K.H.

  Sepiapterin reductase from Chlorobium tepidum (CT-SR) produces L-threo-tetrahydrobiopterin, an isomer of tetrahydrobiopterin, in the last step of de novo synthesis initiating from GTP. Native CT-SR and a selenomethionine (SeMet) derivative of CT-SR have been crystallized by the hanging-drop vapour ... >> More

  Sepiapterin reductase from Chlorobium tepidum (CT-SR) produces L-threo-tetrahydrobiopterin, an isomer of tetrahydrobiopterin, in the last step of de novo synthesis initiating from GTP. Native CT-SR and a selenomethionine (SeMet) derivative of CT-SR have been crystallized by the hanging-drop vapour-diffusion method using PEG 400 as precipitant. CT-SR crystals belong to space group R32, with unit-cell parameters a = b = 201.142, c = 210.184 A, and contain four molecules in the asymmetric unit. Diffraction data were collected to 2.1 A resolution using synchrotron radiation. The structure of CT-SR has been determined using MAD phasing. There is one CT-SR tetramer in the asymmetric unit formed by two closely interacting CT-SR dimers. The solvent content is calculated to be about 67.2%. << Less

  Acta Crystallogr Sect F Struct Biol Cryst Commun 61:202-204(2005) [PubMed] [EuropePMC]

  This publication is cited by 4 other entries.

• Sepiapterin reductase exhibits a NADPH-dependent dicarbonyl reductase activity.

  Katoh S., Sueoka T.

  We have found a new ability of sepiapterin reductase, which has been known to show a strict substrate specificity for the 6-lactyl sidechain of sepiapterin to produce 6-dihydroxypropyl sidechain of dihydrobiopterin in the biosynthesis of tetrahydrobiopterin, to reduce many dicarbonyl compounds wit ... >> More

  We have found a new ability of sepiapterin reductase, which has been known to show a strict substrate specificity for the 6-lactyl sidechain of sepiapterin to produce 6-dihydroxypropyl sidechain of dihydrobiopterin in the biosynthesis of tetrahydrobiopterin, to reduce many dicarbonyl compounds with NADPH as effectively utilized substrates. By analysis of diacetyl reduction by purified sepiapterin reductase, it was observed that both of the carbonyl groups of the compound are finally sequentially reduced by the enzyme with NADPH to hydroxyl groups. And we expect that this enzyme may reduce "Compound X", which is an intermediate of tetrahydrobiopterin synthesis and would be a dicarbonyl derivative of pteridine (Tanaka et. al., 1980), to dihydrobiopterin via sepiapterin. << Less

  Biochem Biophys Res Commun 118:859-866(1984) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Synthesis and characterization of 3H-labelled tetrahydrobiopterin.

  Werner E.R., Schmid M., Werner-Felmayer G., Mayer B., Wachter H.

  We synthesized [3'-3H]-5,6,7,8-tetrahydrobiopterin from [8,5'-3H]guanosine 5'-triphosphate ([8,5'-3H]GTP) using GTP cyclohydrolase (EC 3.5.4.16), 6-pyruvoyltetrahydropterin synthase and sepiapterin reductase (EC 1.1.1.153). After purification by cation-exchange h.p.l.c. a solution of radiochemical ... >> More

  We synthesized [3'-3H]-5,6,7,8-tetrahydrobiopterin from [8,5'-3H]guanosine 5'-triphosphate ([8,5'-3H]GTP) using GTP cyclohydrolase (EC 3.5.4.16), 6-pyruvoyltetrahydropterin synthase and sepiapterin reductase (EC 1.1.1.153). After purification by cation-exchange h.p.l.c. a solution of radiochemically pure (> 95%) [3'-3H]-5,6,7,8-tetrahydrobiopterin with a specific activity of 9.2 Ci/mmol was obtained. The product proved well suited for studying the binding of tetrahydrobiopterin to nitric-oxide synthase. << Less

  Biochem J 304:189-193(1994) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• The 1.25-A crystal structure of sepiapterin reductase reveals its binding mode to pterins and brain neurotransmitters.

  Auerbach G., Herrmann A., Gutlich M., Fischer M., Jacob U., Bacher A., Huber R.

  Sepiapterin reductase catalyses the last steps in the biosynthesis of tetrahydrobiopterin, the essential co-factor of aromatic amino acid hydroxylases and nitric oxide synthases. We have determined the crystal structure of mouse sepiapterin reductase by multiple isomorphous replacement at a resolu ... >> More

  Sepiapterin reductase catalyses the last steps in the biosynthesis of tetrahydrobiopterin, the essential co-factor of aromatic amino acid hydroxylases and nitric oxide synthases. We have determined the crystal structure of mouse sepiapterin reductase by multiple isomorphous replacement at a resolution of 1.25 A in its ternary complex with oxaloacetate and NADP. The homodimeric structure reveals a single-domain alpha/beta-fold with a central four-helix bundle connecting two seven-stranded parallel beta-sheets, each sandwiched between two arrays of three helices. Ternary complexes with the substrate sepiapterin or the product tetrahydrobiopterin were studied. Each subunit contains a specific aspartate anchor (Asp258) for pterin-substrates, which positions the substrate side chain C1'-carbonyl group near Tyr171 OH and NADP C4'N. The catalytic mechanism of SR appears to consist of a NADPH-dependent proton transfer from Tyr171 to the substrate C1' and C2' carbonyl functions accompanied by stereospecific side chain isomerization. Complex structures with the inhibitor N-acetyl serotonin show the indoleamine bound such that both reductase and isomerase activity for pterins is inhibited, but reaction with a variety of carbonyl compounds is possible. The complex structure with N-acetyl serotonin suggests the possibility for a highly specific feedback regulatory mechanism between the formation of indoleamines and pteridines in vivo. << Less

  EMBO J. 16:7219-7230(1997) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• Mutational analysis of sites in sepiapterin reductase phosphorylated by Ca2+/calmodulin-dependent protein kinase II.

  Fujimoto K., Takahashi S.Y., Katoh S.

  Sepiapterin reductase (SPR) catalyzes the last step in the pathway of tetrahydrobiopterin biosynthesis in tissues. SPR is phosphorylated by Ca2+-dependent protein kinases, which indicates that Ca2+-activated protein kinases may play a role in the regulation of SPR in vivo. Phosphorylation sites of ... >> More

  Sepiapterin reductase (SPR) catalyzes the last step in the pathway of tetrahydrobiopterin biosynthesis in tissues. SPR is phosphorylated by Ca2+-dependent protein kinases, which indicates that Ca2+-activated protein kinases may play a role in the regulation of SPR in vivo. Phosphorylation sites of rat sepiapterin reductase (rSPR) by Ca2+/calmodulin-dependent protein kinase II were determined in the present study. Using specific monoclonal anti-phospho-Ser and -Thr antibodies, we found that only Ser residues of rSPR were phosphorylated. We constructed several point mutants of SPR by systematically replacing the three Ser residues by Ala ones. These mutants showed that all three Ser residues, i.e. S46, S196, and S214, of rSPR were phosphorylated. We also recognized that only Ser-213 of human SPR was phosphorylated. Each of these serine residues in SPR was found in the consensus sequence (Arg-X-X-Ser/Thr) of the phosphorylation site. << Less

  Biochim. Biophys. Acta 1594:191-198(2002) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Characterization of recombinant Dictyostelium discoideum sepiapterin reductase expressed in E. coli.

  Kim Y.-A., Chung H.J., Kim Y.J., Choi Y.K., Hwang Y.K., Lee S.W., Park Y.S.

  A cDNA clone (SSC801) putatively encoding sepiapterin reductase (SR) was obtained from the expressed sequence tag clones of Dictyostelium discoideum. The cDNA sequence of 878 nucleotides constituted an ORF of 265 amino acid residues but was missing a few N-terminal residues. The deduced amino acid ... >> More

  A cDNA clone (SSC801) putatively encoding sepiapterin reductase (SR) was obtained from the expressed sequence tag clones of Dictyostelium discoideum. The cDNA sequence of 878 nucleotides constituted an ORF of 265 amino acid residues but was missing a few N-terminal residues. The deduced amino acid sequence showed 29.8% identity with mouse SR sequence and a molecular mass of 29,969 Da. The coding sequence was cloned in E. coli expression vector and overexpressed. The purified His-tag recombinant enzyme was confirmed to have the genuine activity of SR to produce tetrahydrobiopterin from 6-pyruvoyltetrahydropterin in a coupled assay with 6-pyruvoyltetrahydropterin synthase as well as dihydrobiopterin from sepiapterin. However, dictyopterin was not observed in our assay condition. The enzyme was also inhibited by N-acetylserotonin and to a lesser extent by melatonin. Km values for NADPH and sepiapterin were 51.8+/-2.7 microM and 40+/-2 microM, respectively. Vmax was determined as 0.14 micromol/min/mg of protein. << Less

  Mol. Cells 10:405-410(2000) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Structure of Chlorobium tepidum sepiapterin reductase complex reveals the novel substrate binding mode for stereospecific production of L-threo-tetrahydrobiopterin.

  Supangat S., Seo K.H., Choi Y.K., Park Y.S., Son D., Han C.D., Lee K.H.

  Sepiapterin reductase (SR) is involved in the last step of tetrahydrobiopterin (BH(4)) biosynthesis by reducing the di-keto group of 6-pyruvoyl tetrahydropterin. Chlorobium tepidum SR (cSR) generates a distinct BH(4) product, L-threo-BH(4) (6R-(1'S,2'S)-5,6,7,8-BH(4)), whereas animal enzymes produ ... >> More

  Sepiapterin reductase (SR) is involved in the last step of tetrahydrobiopterin (BH(4)) biosynthesis by reducing the di-keto group of 6-pyruvoyl tetrahydropterin. Chlorobium tepidum SR (cSR) generates a distinct BH(4) product, L-threo-BH(4) (6R-(1'S,2'S)-5,6,7,8-BH(4)), whereas animal enzymes produce L-erythro-BH(4) (6R-(1'R,2'S)-5,6,7,8-BH(4)) although it has high amino acid sequence similarities to the other animal enzymes. To elucidate the structural basis for the different reaction stereospecificities, we have determined the three-dimensional structures of cSR alone and complexed with NADP and sepiapterin at 2.1 and 1.7 A resolution, respectively. The overall folding of the cSR, the binding site for the cofactor NADP(H), and the positions of active site residues were quite similar to the mouse and the human SR. However, significant differences were found in the substrate binding region of the cSR. In comparison to the mouse SR complex, the sepiapterin in the cSR is rotated about 180 degrees around the active site and bound between two aromatic side chains of Trp-196 and Phe-99 so that its pterin ring is shifted to the opposite side, but its side chain position is not changed. The swiveled sepiapterin binding results in the conversion of the side chain configuration, exposing the opposite face for hydride transfer from NADPH. The different sepiapterin binding mode within the conserved catalytic architecture presents a novel strategy of switching the reaction stereospecificities in the same protein fold. << Less

  J. Biol. Chem. 281:2249-2256(2006) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Carbonyl reductase activity of sepiapterin reductase from rat erythrocytes.

  Sueoka T., Katoh S.

  A homogeneous preparation of sepiapterin reductase, an enzyme involved in the biosynthesis of tetrahydrobiopterin, from rat erythrocytes was found to be responsible for the reduction with NADPH of various carbonyl compounds of non-pteridine derivatives including some vicinal dicarbonyl compounds w ... >> More

  A homogeneous preparation of sepiapterin reductase, an enzyme involved in the biosynthesis of tetrahydrobiopterin, from rat erythrocytes was found to be responsible for the reduction with NADPH of various carbonyl compounds of non-pteridine derivatives including some vicinal dicarbonyl compounds which were reported in the previous paper (Katoh, S. and Sueoka, T. (1984) Biochem, Biophys. Res. Commun. 118, 859-866) in addition to the general substrate, sepiapterin (2-amino-4-hydroxy-6-lactoyl-7,8-dihydropteridine). The compounds sensitive as substrates of the enzyme were quinones, e.g., p-quinone and menadione; other vicinal dicarbonyls, e.g., methylglyoxal and phenylglyoxal; monoaldehydes, e.g., p-nitrobenzaldehyde; and monoketones, e.g., acetophenone, acetoin, propiophenone and benzylacetone. Rutin, dicoumarol, indomethacin, and ethacrynic acid inhibited the enzyme activity toward either a carbonyl compound of a non-pteridine derivative or sepiapterin as substrate. Sepiapterin reductase is quite similar to general aldo-keto reductases, especially to carbonyl reductase. << Less

  Biochim Biophys Acta 843:193-198(1985) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.