SYNTHESIS OF <scp>d</scp>-FRUCTOPYRANOSE 2-PHOSPHATE AND <scp>d</scp>-FRUCTOFURANOSE 2-PHOSPHATE

Pontis, Horacio G.; Fischer, Catalina L.

doi:10.1042/bj0890452

Cited by 39 publications

(15 citation statements)

References 17 publications

(15 reference statements)

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“…2) were acidified to pH 1 -2 and then assayed for fructose. According to Pontis and Fisher [15], who first described the synthesis and characterization of Fru2P, lability of the phosphate group in Fru2P at pH 5 is characteristic for this sugar phosphate. The 6-phosphate group in Fru6P, as well as in Fru(2,6)Pz, is not acid-labile.…”

Section: Resultsmentioning

confidence: 99%

“…3 ) edge on the properties of fructose derivatives, there is no product known other than Fru2P that forms free fructose and inorganic phosphate upon mild acidification. (b) The product that liberates fructose and phosphate upon treatment with mild acid coelutes on ion-exchange chromatography on Dowex 1 with authentic synthetic Fru2P [15].…”

Section: Resultsmentioning

confidence: 99%

“…The eluted fractions were analyzed for sugars with the orcinol reaction [16] and for inorganic phosphate according to Ames [17]. The elution profile of chromatography of the chemical preparation of Fru2P [15] on the Dowex 1 column used in Fig. 3 exhibits peaks of total hexose and of phosphate at the same positions as with the enzymatic incubation (data not shown).…”

Section: Resultsmentioning

confidence: 99%

“…The chemical synthesis of Fru2P was performed as described by Pontis and Fischer [15]. Total carbohydrate was assayed by the orcinol reaction [I61 and orthophosphate by the method of Ames [17].…”

Section: Enzyme Assaysmentioning

confidence: 99%

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Fructose 2-phosphate, an intermediate of the dephosphorylation of fructose 2,6-bisphosphate with a purified yeast enzyme

1987

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A fructose-2,6-bisphosphate dephosphorylating enzyme was 3000-fold purified to electrophoretic homogeneity from Saccharomyces cerevisiae. Half-maximal activity was obtained at pH 6.0 with 6 pM fructose 2,6-bisphosphate and 0.15mM Mg2+. On incubation for 90min with fructose 2,6-bisphosphate, about 80% of the substrate appears with an almost linear time dependence as fructose. In the first 30 min a substance accumulates to about 40% of the consumed fructose 2,6-bisphosphate which forms free fructose on mild acid treatment. Formation of fructose 6-phosphate was negligible. The mild-acid-labile intermediate was identified as fructose 2-phosphate by comparative ion-exchange chromatography with authentic fructose 2-phosphate synthesized from fructose l-phosphate [Pontis, H. . The data suggest the reaction sequence fructose 2,6-bisphosphate + fructose 2-phosphate -+ fructose. The designation fructose-2,6-bisphosphate 6-phosphohydrolase is proposed for the enzyme described here.Fructose 2,6-bisphosphate [Fru(2,6)P2] has' been discovered to be a potent activator of 6-phosphofructokinase-1 and inhibitor of fructose-l,6-bisphosphatase in liver, other mammalian tissues and yeast [l, 21. Fru(2,6)P2 is formed in liver by an enzyme catalyzing ATP-dependent phosphorylation of Fru6P in the 2-position. It has been shown that the same enzyme protein also catalyzes dephosphorylation of Fru(2,6)P2 to Fru6P and inorganic phosphate. Modification of the kinase activity to the phosphatase activity of this enzyme is effected by cyclic-AMP-dependent protein phosphorylation. The reverse modification is the result of dephosphorylation of the enzyme [3, 41. In yeast cells a kinase catalyzing phosphorylation of Fru6P to Fru(2,6)P2 has also been described [5, 61. The kinase is activated by cyclic-AMPdependent phosphorylation [5, 71. This is in contrast to liver, where cyclic-AMP-dependent phosphorylation inactivates fructose-6-phosphate kinase-2 and increases fructose-2,6-bisphosphatase activity. In yeast no activity dephosphorylating Fru(2,6)P2 to Fru6P has been detected [5, 71. It is evident from these results that the metabolism of Fru(2,6)P2 in yeast is different from that in mammalian tissues. In the present paper purification of an enzyme to electrophoretic homogeneity from yeast which dephosphorylates Fru(2,6)P2 is described. Fru2P, which has not been described previously as an enzymatic product, and fructose were shown to be the dephosphorylation products.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Enzyme Assaysmentioning

confidence: 99%

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Fructose 2-phosphate, an intermediate of the dephosphorylation of fructose 2,6-bisphosphate with a purified yeast enzyme

1987

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“…With respect to the position of the phosphorylated carbon in DMF-phosphate, tandem MS allowed us to exclude carbons C4-C6, suggesting that either C2 or C3 are phosphorylated. However, C2 seems unlikely as furanosyl-phosphates, such as fructose 2-phosphate [27] and fructose 2,6-bisphosphate [28], are extremely labile in acid even in the cold ; a 2-phosphate derivative would have therefore been hydrolysed during the extraction with perchloric acid. Furthermore, the NMR spectra show several anomeric forms of the fructosamine-phosphate derivative, which indicates that the carbohydrate moiety is free to anomerize and therefore that the hydroxy group on C2 of the 1-deoxyfructose moiety is free.…”

Section: Discussionmentioning

confidence: 99%

Conversion of a synthetic fructosamine into its 3-phospho derivative in human erythrocytes

Delpierre

Vanstapel

Stroobant

et al. 2000

Biochemical Journal

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Intact human erythrocytes catalyse the conversion of fructose into fructose 3-phosphate with an apparent Km of 30mM [Petersen, Kappler, Szwergold and Brown (1992) Biochem. J. 284, 363Ő366]. The physiological significance of this process is still unknown. In the present study we report that the formation of fructose 3-phosphate from 50mM fructose in intact erythrocytes is inhibited by 1-deoxy-1-morpholinofructose (DMF), a synthetic fructosamine, with an apparent Ki of 100µM. 31P NMR analysis of cell extracts incubated with DMF indicated the presence of an additional phosphorylated compound, which was partially purified and shown to be DMF 3-phosphate by tandem MS. Radiolabelled DMF was phosphorylated by intact erythrocytes with an apparent Km (≈ 100µM) approx. 300-fold lower than the value reported for fructose phosphorylation on its third carbon. These results indicate that the physiological function of the enzyme that is able to convert fructose into fructose 3-phosphate in intact erythrocytes is probably to phosphorylate fructosamines. This suggests that fructosamines, which are produced non-enzymically from glucose and amino compounds, may be metabolized in human erythrocytes.

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Fructose 2,6‐Bisphosphate

Schaftingen

1987

Advances in Enzymology - And Related Areas of Molecular Biology

170

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SYNTHESIS OF d-FRUCTOPYRANOSE 2-PHOSPHATE AND d-FRUCTOFURANOSE 2-PHOSPHATE

Abstract: The cyclic phosphates were already well separated after 8-10 hr., but a longer run is convenient for obtaining better resolutions between the other D-fructose phosphates.Vol. 89

Cited by 39 publications

References 17 publications

Fructose 2-phosphate, an intermediate of the dephosphorylation of fructose 2,6-bisphosphate with a purified yeast enzyme

Fructose 2-phosphate, an intermediate of the dephosphorylation of fructose 2,6-bisphosphate with a purified yeast enzyme

Conversion of a synthetic fructosamine into its 3-phospho derivative in human erythrocytes

Fructose 2,6‐Bisphosphate

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