Abstract
Although fructans occur widely in several plant families and they have been a subject of investigation for decennia, the mechanism of their biosynthesis is not completely elucidated. We succeeded in purifying a fructan: fructan 1-fructosyl transferase (1-FFT; EC 2.4.1.100) from chicory roots (Cichorium intybus L. var. foliosum cv. Flash). In combination with the purified chicory root sucrose: sucrose 1-fructosyl transferase (1-SST; EC 2.4.1.99), this enzyme synthesized a range of naturally occurring chicory fructans (inulins) from sucrose as the sole substrate. Starting from physiologically relevant sucrose concentrations, inulins up to a degree of polymerization (DP) of about 20 were synthesized in vitro after 96 h at 0°C. Neither 1-SST, nor 1-FFT alone could mediate the observed fructan synthesis. Fructan synthesis in vitro was compared starting from 50, 100 and 200 mM sucrose, respectively. The initiation of (DP > 3)-fructan synthesis was found to be correlated with a certain ratio of 1 kestose to sucrose. The data presented now provide strong evidence to validate the 1-SST/1-FFT model for in-vivo fructan synthesis, at least in the Asteraceae.
Similar content being viewed by others
Abbreviations
- DP:
-
degree of polymerization
- 1-FFT:
-
fructan: fructan 1-fructosyl transferase
- 1-SST:
-
sucrose: sucrose 1-fructosyl transferase
References
Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248–254
Cairns AJ (1993) Evidence for the de novo synthesis of fructan by enzymes from higher plants: a reappraisal of the 1-SST/1-FFT model. New Phytol 123: 15–24
Cairns AJ, Ashton JE (1991) The interpretation of in vitro measurements of fructosyl transferase activity: an analysis of patterns of fructosyl transfer by fungal invertase. New Phytol 118: 23–34
Cairns AJ, Ashton JE (1993) Species-dependent patterns of fructan synthesis by enzymes from excised leaves of oat, wheat, barley and timothy. New Phytol 124: 381–388
Cairns AJ, Ashton JE (1994) Fructan biosynthesis in excised leaves of Lolium temulentum L. VI. Optimization and stability of enzymatic fructan synthesis. New Phytol 126: 3–10
Chatterton NJ, Harrison PA, Thornley WR, Bennett JH (1993) Separation and quantification of fructan (inulin) oligomers by anion exchange chromatography. In: Fuchs A (ed) Inulin and inulin-containing crops. Elsevier, Amsterdam, pp 93–99
Chen JQ, Black CC (1992) Biochemical and immunological properties of alkaline invertase isolated from sprouting soybean hypocotyls. Arch Biochem Biophys 2951: 61–69
Darwen CWE, John P (1989) Localisation of the enzymes of fructan metabolism in vacuoles isolated by a mechanical method from tubers of Jerusalem artichoke (Helianthus tuberosus L.). Plant Physiol 89: 658–663
Edelman J, Jefford TG (1968) The mechanism of fructan metabolism in higher plants as exemplified in Helianthus tuberosus. New Phytol 67: 517–531
Fuchs A (1991) Current and potential food and non-food applications of fructans. Biochem Soc Trans 19: 555–560
Hendry G (1993) Evolutionary origins and natural functions of fructans. A climatological, biogeographic and mechanistic appraisal. New Phytol 123: 3–14
Hidaka H, Hirayama M, Yamada K (1991) Fructooligosaccharides. Enzymatic preparation and biofunctions. J Carbohydr Chem 10: 509–522
Meier H, Reid JSG (1982) Reserve polysaccharides other than starch in higher plants. In: Loewus FA, Tanner W (eds) Encyclopedia of plant physiology (NS), Plant carbohydrates. Springer-Verlag, Berlin, pp 418–471
Penson SP, Cairns AJ (1994) Fructan biosynthesis in excised leaves of wheat (Triticum aestivum L.): a comparison of de novo synthesis in vivo and in vitro. New Phytol 128: 395–402
Shiomi N, Onodera S, Chatterton NJ, Harrison PA (1991) Separation of fructo-oligosaccharide isomers by anion-exchange chromatography. Agric Biol Chem 55: 1427–1428
Van den Ende W, Van Laere A (1993) Purification and properties of an invertase with sucrose: sucrose fructosyltransferase (1-SST) activity from the roots of Cichorium intybus L. New Phytol 123: 31–37
Van den Ende W, Van Laere A (1995) Purification and properties of a neutral invertase from the roots of Cichorium intybus. Physiol Plant 93: 241–248
Van den Ende W, Van Wonterghem D, Verhaert P, Dewil E, Van Laere A (1996a) Purification and characterization of fructan: fructan fructosyl transferase from chicory roots (Cichorium intybus L.). Planta, in press
Van den Ende W, Van Wonterghem D, Verhaert P, Dewil E, De Loof A, Van Laere A (1996b) Purification and characterization of 1-SST, the key enzyme initiating fructan biosynthesis in young chicory roots (Cichorium intybus L.). Physiologia Plantarum, in press
Wiemken A, Frehner M, Keller F, Wagner W (1986) Fructan metabolism, enzymology and compartmentation. Curr Top Plant Biochem Physiol 5: 17–37
Wiemken A, Sprenger N, Boller T (1995) Fructans — an extension of sucrose by sucrose. In: Pontis HG, Salerno GL, Echeverria EJ (eds) Current topics in plant physiology, vol 14: Sucrose metabolism, biochemistry, physiology and molecular biology. ASPP, Rockville, pp 179–189
Author information
Authors and Affiliations
Additional information
The authors thank E. Nackaerts for valuable technical assistance. W. Van den Ende is grateful to the National Fund for Scientific Research (NFSR Belgium) for giving a grant for research assistants.
Rights and permissions
About this article
Cite this article
Van den Ende, W., Van Laere, A. De-novo synthesis of fructans from sucrose in vitro by a combination of two purified enzymes (sucrose: sucrose 1-fructosyl transferase and fructan: fructan 1-fructosyl transferase) from chicory roots (Cichorium intybus L.). Planta 200, 335–342 (1996). https://doi.org/10.1007/BF00200301
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00200301