Purification and Characterization of Wheat β(2→1) Fructan:Fructan Fructosyl Transferase Activity

Abstract

Fructans are the major storage carbohydrate in vegetative tissues of wheat (Triticum aestivum L.). Fructan:fructan fructosyl transferase (FFT) catalyzes fructosyl transfer between fructan molecules to elongate the fructan chain. The objective of this research was to isolate this activity in wheat. Wheat (cv Caldwell) plants grown at 25°C for 3 weeks were transferred to 10°C to induce fructan synthesis. From the leaf blades kept at 10°C for 4 days, fructosyl transferase activity was purified using salt precipitation and a series of chromatographic procedures including size exclusion, anion-exchange, and affinity chromatography. The transferase activity was free from invertase and other fructan-metabolizing activities. Fructosyl transferase had a broad pH spectrum with a peak activity at 6.5. The temperature optimum was 30°C. The activity was specific for fructosyl transfer from β(2→1)-linked 1-kestose or fructan to sucrose and β(2→1) fructosyl transfer to other fructans (1-FFT). Fructosyl transfer from oligofructans to sucrose was most efficient when 1-kestose was used as donor molecule and declined as the degree of polymerization of the donor increased from 3 to 5. 1-FFT catalyzed the in vitro synthesis of inulin tetra- and penta-saccharides from 1-kestose; however, formation of the tetrasaccharide was greatly reduced at high sucrose concentration. 6-Kestose could not act as donor molecule, but could accept a fructosyl moiety from 1-kestose to produce bifurcose and a tetrasaccharide having a β(2→1) fructose attached to the terminal fructose of 6-kestose. The role of this FFT activity in the synthesis of fructan in wheat is discussed.