Development of high-affinity ligands and photoaffinity labels for the D-fructose transporter GLUT5

Jing Yang, James Dowden, Arnaud Tatibouët, Yasumaru Hatanaka, Geoffrey D. Holman

Research output: Contribution to journalArticle

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Abstract

The GLUT5 transporter catalyses the specific uptake Of D-fructose and can accept this hexose in its furanose and pyranose ring forms. The transporter does not accept fructose epimers and has very limited tolerance of bulky groups substituted at the 2-, 3-, 4- and 5-OH positions [Tatibouet, Yang, Morin and Holman (2000) Bioorg. Med. Chem. 8, 1825-1833]. To further explore whether bulky groups can be tolerated at the primary OH positions, a D-fructose analogue with an allylamine group substitution to replace the 1-OH group was synthesized and was found to be quite well tolerated (K-i = 27.1 mM). However, this analogue occurs in multiple ring forms. By contrast, 2,5-anhydro-D-mannitol is a symmetrical molecule that occurs only in a furanose ring form in which C-1 and C-6 are equivalent. We have therefore synthesized new 2,5-anhydro-D-mannitol analogues (substituted at the equivalent of the 6-OH Of D-fructose) and from studies in Chinese hamster ovary cells expressing GLUT5 cells report that (i) the allylamine derivative of 2,5-anhydro-D-mannitol is well tolerated (K-i = 2.66 mM); (ii) introduction of a di-nitrophenyl-substituted secondary amine group enhances affinity (K-i = 0.56 mM); (iii) introduction of amide-linked biotinylated photolabel moieties is possible without loss of affinity relative to 2,5-anhydro-D-mannitol but a small secondary amine spacer between the biotinylated photolabelling moiety and the fructofuranose ring increases affinity (fructose photolabel 2; A = 1.16 mM); (iv) introduction of a hydrophilic tartarate spacer between biotin and the diazirine photoreactive groups can be accomplished without reduction in affinity and (v) photoactivation of biotinylated fructose photolabels leads to specific biotin tagging of GLUT5. These data suggest that substitution of a secondary amine group (-NH) to replace the C-6 (or C-1) -OH of 2,5-anhydro-D-mannitol results in compounds of high affinity; the affinity is enhanced over 10-fold compared with D-fructose.
Original languageEnglish
Pages (from-to)533-539
Number of pages7
JournalBiochemical Journal
Volume367
Issue number2
DOIs
Publication statusPublished - 2002

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Glucose Transporter Type 5
Photoaffinity Labels
Affinity Labels
Fructose
Ligands
Allylamine
Amines
Biotin
Substitution reactions
Diazomethane
Hexoses
Cricetulus
Amides
Ovary
Cells
2,5-anhydromannitol
Derivatives

Cite this

Development of high-affinity ligands and photoaffinity labels for the D-fructose transporter GLUT5. / Yang, Jing; Dowden, James; Tatibouët, Arnaud; Hatanaka, Yasumaru; Holman, Geoffrey D.

In: Biochemical Journal, Vol. 367, No. 2, 2002, p. 533-539.

Research output: Contribution to journalArticle

Yang, Jing ; Dowden, James ; Tatibouët, Arnaud ; Hatanaka, Yasumaru ; Holman, Geoffrey D. / Development of high-affinity ligands and photoaffinity labels for the D-fructose transporter GLUT5. In: Biochemical Journal. 2002 ; Vol. 367, No. 2. pp. 533-539.
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T1 - Development of high-affinity ligands and photoaffinity labels for the D-fructose transporter GLUT5

AU - Yang, Jing

AU - Dowden, James

AU - Tatibouët, Arnaud

AU - Hatanaka, Yasumaru

AU - Holman, Geoffrey D.

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PY - 2002

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N2 - The GLUT5 transporter catalyses the specific uptake Of D-fructose and can accept this hexose in its furanose and pyranose ring forms. The transporter does not accept fructose epimers and has very limited tolerance of bulky groups substituted at the 2-, 3-, 4- and 5-OH positions [Tatibouet, Yang, Morin and Holman (2000) Bioorg. Med. Chem. 8, 1825-1833]. To further explore whether bulky groups can be tolerated at the primary OH positions, a D-fructose analogue with an allylamine group substitution to replace the 1-OH group was synthesized and was found to be quite well tolerated (K-i = 27.1 mM). However, this analogue occurs in multiple ring forms. By contrast, 2,5-anhydro-D-mannitol is a symmetrical molecule that occurs only in a furanose ring form in which C-1 and C-6 are equivalent. We have therefore synthesized new 2,5-anhydro-D-mannitol analogues (substituted at the equivalent of the 6-OH Of D-fructose) and from studies in Chinese hamster ovary cells expressing GLUT5 cells report that (i) the allylamine derivative of 2,5-anhydro-D-mannitol is well tolerated (K-i = 2.66 mM); (ii) introduction of a di-nitrophenyl-substituted secondary amine group enhances affinity (K-i = 0.56 mM); (iii) introduction of amide-linked biotinylated photolabel moieties is possible without loss of affinity relative to 2,5-anhydro-D-mannitol but a small secondary amine spacer between the biotinylated photolabelling moiety and the fructofuranose ring increases affinity (fructose photolabel 2; A = 1.16 mM); (iv) introduction of a hydrophilic tartarate spacer between biotin and the diazirine photoreactive groups can be accomplished without reduction in affinity and (v) photoactivation of biotinylated fructose photolabels leads to specific biotin tagging of GLUT5. These data suggest that substitution of a secondary amine group (-NH) to replace the C-6 (or C-1) -OH of 2,5-anhydro-D-mannitol results in compounds of high affinity; the affinity is enhanced over 10-fold compared with D-fructose.

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