Fructan biosynthesis in excised leaves of Lolium temulentum L.: V. Enzymatic de novo synthesis of large fructans from sucrose.

Leaves of Lolium temulentum L. were induced to accumulate fructan by excision and continuous illumination. Enzyme extracts were prepared and protein was concentrated by precipitation with ammonium sulphate at 100% saturation. The preparation was incubated at a protein concentration equivalent to 3-4 g f.wt of tissue cm with sucrose at an initial concentration of 400 mol . The preparation catalyzed the synthesis of large fructans of apparent degree of polymerization (DP) ⩽20, based upon comparison with the chromatographic mobilities of oligoinulins on thin-layer chromatography (TLC). The products were formed at a rate approximating to rates of fructan accumulation in leaf tissue. The overall pattern of in vitro products, when separated on TLC, resembled that of the native leaf fructans though the relative abundances of some of the products differed from the in vivo pattern. The reaction did not catalyze the formation of the high molecular weight fructans (M > 3 kDa; DP > 20) characteristic of this tissue. Equivalent protein preparations derived from uninduced leaves exhibited negligible rates of fructan synthesis. Feeding cycloheximide to leaves prevented fructan synthesis both by the tissue and by protein extracts. Cycloheximide did not directly inhibit in vitro fructan synthesis, suggesting that its in vivo effect was at the translational level. The characteristics of the in vitro reaction are compared with the properties of in vivo fructan synthesis and are discussed with respect to its possible physiological relevance.