Fructan exohydrolase from grasses
Authors: SIMPSON RICHARD J. , BONNETT GRAHAM D. 
Published: April 2006

In grasses, fructan reserves are mobilized from vegetative plant parts during seasonal growth, after defoliation during grazing and from stems during seed filling. Well‐illuminated leaves show a diurnal pattern of fructan accumulation during the light and mobilization during the dark. In expanding leaves, fructans are accumulated in cells of the elongation zone and when mobilized are considered to contribute assimilate for synthetic processes. Even in leaves which do not contain high fructan concentrations, high rates of fructan turnover occur. The process of fructan mobilization appears to be regulated in relation to ontogenic events, demand for assimilate during growth and in response to environmental stress.

Hydrolysis of fructans in bacteria is catalyzed by both endo‐ and exohydrolases. However, in higher plants only fructan exohydrolases (FEH) (EC 3.2.1.80) have been reported. FEH has been extracted from only a limited number of grass species. The pH optimum of FEH activities varies between pH 45‐5‐5, the temperature optimum ranges from 25–40 °C and FEH is considered to be entirely localized in vacuoles. Estimates of the Km for FEH assayed using high molecular weight fructan substrates vary widely and should be considered carefully because most substrates are ill‐defined. Many studies indicate that crude and partially‐purified FEH activity is highest when assayed using a fructan substrate extracted from the species that was the source of the enzyme activity. Inulin extracted from members of the Asteraceae is generally less readily hydrolyzed and levans from bacteria are relatively poor substrates for FEH from grasses. Glycosidic‐linkage‐specific hydrolysis has been demonstrated for an FEH activity extracted from barley. This FEH activity hydrolyzed β‐2,1‐glycosidic linkages more rapidly than β‐2,6‐linkages. Most other studies are less conclusive because ill‐defined fructan substrates were used. Two isoforms of FEH are reported in leaves of Lolium spp., but the roles of isoforms and their kinetic characteristics are not known.

FEH activity in different tissues may be regulated by metabolic concentrations, sucrose (5–10 mw) being a strong inhibitor in vitro of FEH from some species. Results of experiments with Dactylis glomerata indicate control of expression of FEH activity at the gene level. In stem bases, FEH activity increased after defoliation. The increase was abolished by applications of inhibitors of protein synthesis and was apparently repressed by application of various sugars.

Although the rates of fructan hydrolysis measured in vitro are sufficient to explain the in vivo rates of fructan hydrolysis, it is yet to be shown whether fructan hydrolysis in vivo is due to the activity of FEH exclusively, or FEH and invertase‐like activities.

The overriding conclusion is that the various studies of FEH from grasses present a confusing and incomplete picture of the function, activity and kinetics of this enzyme. This is due in part to the lack of defined, commercially‐available substrates. The chromatographic techniques available to most laboratories do not permit purification of sufficient quantities of high molecular weight fructans of specific degree of polymerization, or fructan oligosaccharides with glycosidic linkages which differ from that of the inulin series for enzyme characterization. It is recommended that a few well‐defined oligosaccharides be adopted as substrate standards for future research.