Fructan metabolism in wheat (Triticum aestivum L.)
Fructans are the major vegetative storage carbohydrate in wheat (Triticum aestivum L.). The objectives of this study were (1) to develop a system in which the direction of fructan metabolism could be shifted or controlled, (2) to isolate and characterize fructan exohydrolase (FEH) and fructan:fructan fructosyl transferase (FFT). Three-week-old wheat seedlings (cv. Caldwell) grown at 25$\sp\circ$C transferred to 10$\sp\circ$C to induce fructan synthesis and after leaf blades excision at day 4 plants transferred to darkness at 25$\sp\circ$C. Fructan content increased significantly one day after transfer to 10$\sp\circ$C in leaf blades and sheaths (90% sheaths and 10% stem tissue) with a greater concentration of high molecular weight fructan in sheaths. Sheath fructan content declined rapidly following transfer to 25$\sp\circ$C in darkness. Invertase activity in leaf blades fluctuated during cold treatment, while sheath invertase activity decreased and remained low. In leaf blades and sheaths sucrose:sucrose fructosyl transferase (SST) activity increased during cold treatment, while FEH activity decreased. Sheath invertase and FEH activity increased after transfer to darkness at 25$\sp\circ$C, while SST activity decreased. FEH was purified 356-fold from stems and sheaths of field-grown wheat (cv. Clark). FEH had a molecular weight of 63.7 kD and an optima at pH 5.5 and 30$\sp\circ$C. The Kms for (2 $\to$ 1) linked oligofructans varied, from 10 to 37 mM, with the lowest Km for tetrasaccharide. The Vmax increased as degree of polymerization (DP) increased. Wheat FEH hydrolyzed only (2 $\to$ 1) linked fructans but not Timothy fructan or sucrose. FFT purified from cold-induced wheat leaves had an optima at pH 6.5 and 30$\sp\circ$C. FFT catalyzed fructosyl transfer from (2 $\to$ 1) linked fructans to sucrose and other fructans. Fructosyl transfer from oligofructans to sucrose as most efficient when 1-kestose was used as donor molecule and declined as the donor DP increased from DP 3 to 5. 6-Kestose could not act as fructosyl donor but could accept a moiety from 1-kestose producing bifurcose. FFT catalyzed synthesis of nystose and inulin DP5 from 1-kestose. However, nystose formation was inhibited by sucrose.
Housley, Purdue University.
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