Biosynthesis, Nonenzymatic Synthesis, and Purification of the Intermediate in Synthesis of Sepiapterin in Drosophila

Dale Dorsett, John Flanagan, K. Bruce Jacobson

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

The enzymatic conversion of the D-erythro-dihydroneopterin triphosphate [H2-neopterin-(P)3] to sepiapterin occurs via a nonphosphorylated intermediate as shown by others. We have developed a high-performance liquid chromatography assay for this intermediate and have found that the intermediate (X) and two related compounds (X1 and X2) can be formed nonenzymatically under certain conditions from H2-neopterin-(P)3. The reaction is catalyzed by tris(hydroxymethyl) aminomethane, dependent upon H2-neopterin-(P)3 concentration, significant at temperatures greater than 80°C, and maximal between pH 8.5 and 9.5 (as determined at 25 °C). All three compounds were purified, and it was found that both X and XI can serve as substrates for the enzymatic, NADPH-dependent synthesis of sepiapterin. From the kinetics of formation from H2-neopterin-(P)3 and the similarity of the ultraviolet spectra, it is clear that X, XI, and X2 are closely related compounds. None of the three compounds is reduced by NaBH4; only XI is sensitive to periodate oxidation. All three can be oxidized with iodine to give rise to highly fluorescent compounds that in turn can be reduced by NaBH4 to give rise to the respective parent compounds. These latter observations indicate that X, XI, and X2 are dihydropterins. These results are discussed relative to the proposed structures for enzymatically produced X. The methods described for the nonenzymatic synthesis of X and its purification should allow preparation of large amounts of X for future study.

Original languageEnglish (US)
Pages (from-to)3892-3899
Number of pages8
JournalBiochemistry
Volume21
Issue number16
DOIs
StatePublished - Jan 1 1982

Fingerprint

Neopterin
Biosynthesis
Drosophila
Purification
Tromethamine
High performance liquid chromatography
NADP
Iodine
Assays
High Pressure Liquid Chromatography
Oxidation
Kinetics
Temperature
sepiapterin
Substrates

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

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title = "Biosynthesis, Nonenzymatic Synthesis, and Purification of the Intermediate in Synthesis of Sepiapterin in Drosophila",
abstract = "The enzymatic conversion of the D-erythro-dihydroneopterin triphosphate [H2-neopterin-(P)3] to sepiapterin occurs via a nonphosphorylated intermediate as shown by others. We have developed a high-performance liquid chromatography assay for this intermediate and have found that the intermediate (X) and two related compounds (X1 and X2) can be formed nonenzymatically under certain conditions from H2-neopterin-(P)3. The reaction is catalyzed by tris(hydroxymethyl) aminomethane, dependent upon H2-neopterin-(P)3 concentration, significant at temperatures greater than 80°C, and maximal between pH 8.5 and 9.5 (as determined at 25 °C). All three compounds were purified, and it was found that both X and XI can serve as substrates for the enzymatic, NADPH-dependent synthesis of sepiapterin. From the kinetics of formation from H2-neopterin-(P)3 and the similarity of the ultraviolet spectra, it is clear that X, XI, and X2 are closely related compounds. None of the three compounds is reduced by NaBH4; only XI is sensitive to periodate oxidation. All three can be oxidized with iodine to give rise to highly fluorescent compounds that in turn can be reduced by NaBH4 to give rise to the respective parent compounds. These latter observations indicate that X, XI, and X2 are dihydropterins. These results are discussed relative to the proposed structures for enzymatically produced X. The methods described for the nonenzymatic synthesis of X and its purification should allow preparation of large amounts of X for future study.",
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Biosynthesis, Nonenzymatic Synthesis, and Purification of the Intermediate in Synthesis of Sepiapterin in Drosophila. / Dorsett, Dale; Flanagan, John; Jacobson, K. Bruce.

In: Biochemistry, Vol. 21, No. 16, 01.01.1982, p. 3892-3899.

Research output: Contribution to journalArticle

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AB - The enzymatic conversion of the D-erythro-dihydroneopterin triphosphate [H2-neopterin-(P)3] to sepiapterin occurs via a nonphosphorylated intermediate as shown by others. We have developed a high-performance liquid chromatography assay for this intermediate and have found that the intermediate (X) and two related compounds (X1 and X2) can be formed nonenzymatically under certain conditions from H2-neopterin-(P)3. The reaction is catalyzed by tris(hydroxymethyl) aminomethane, dependent upon H2-neopterin-(P)3 concentration, significant at temperatures greater than 80°C, and maximal between pH 8.5 and 9.5 (as determined at 25 °C). All three compounds were purified, and it was found that both X and XI can serve as substrates for the enzymatic, NADPH-dependent synthesis of sepiapterin. From the kinetics of formation from H2-neopterin-(P)3 and the similarity of the ultraviolet spectra, it is clear that X, XI, and X2 are closely related compounds. None of the three compounds is reduced by NaBH4; only XI is sensitive to periodate oxidation. All three can be oxidized with iodine to give rise to highly fluorescent compounds that in turn can be reduced by NaBH4 to give rise to the respective parent compounds. These latter observations indicate that X, XI, and X2 are dihydropterins. These results are discussed relative to the proposed structures for enzymatically produced X. The methods described for the nonenzymatic synthesis of X and its purification should allow preparation of large amounts of X for future study.

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