β-Furfuryl-β-Glucoside: An Endogenous Activator of Higher Plant UDP-Glucose:(1-3)-β-Glucan Synthase

Abstract

In a recent paper (P Ohana, DP Delmer, JC Steffens, DE Matthews, R Mayer, M Benziman [1991] J Biol Chem 266: 13472-13475), we described the purification and structural characterization of j@-furfuryl-i-glucoside (FG), an endogenous activator of plant UDP-glucose:(1-+3)-#-glucan (callose) synthase. In the present report, we provide evidence that FG specifically stimulates callose synthase. The effects of FG on the kinetic properties of callose synthase were studied, and we ascertained that FG, or at least a very similar compound, is present in other plant systems. Chemically synthesized a-furfuryl-P-glucoside also stimulates callose synthase, exhibiting a slightiy higher K. of 80 micromolar, compared with 50 micromolar for FG. In addition, we have identified and partially characterized an enzyme that catalyzes the synthesis of FG using 0-furfuryl alcohol and UDP-glucose as substrates. A model for the regulation of callose synthesis in vivo, involving changes in intracellular compartmentation of FG and Ca2+, is proposed.It is well established that the plasma membrane-localized UDP-glucose:(l->3)-}-glucan synthase (callose synthase) has an absolute requirement for Ca2", and is also stimulated by simple (l-glucosides, at high millimolar concentrations (6,12).Kinetic studies have shown that the enzyme is virtually inactive unless both Ca2" and a ,B-glucoside are supplied as effectors. We recently reported the discovery of a unique endogenous activator of plant callose synthase and have defined its precise structure as FG2 (for structure, see Fig. 3 MATERIALS AND METHODS MaterialsSeeds from mung beans (Vigna radiata), sorghum (Sorghum vulgare), barley (Hordeum vulgare), or peas (Pisum sativum) were allowed to germinate on moist filter paper in Petri plates kept in the dark at 25°C. When seedlings reached a length of 2 to 3 cm, roots and seeds were discarded and the remaining tissue was harvested. Cotton fibers (Gossypium hirsutum Acala SJ-2) were obtained from boils harvested 19 to 22 d postanthesis from field-grown plants (7). Prior to extraction, the fibers were ground to a fine powder in liquid N2. Purified native and synthetic FG were obtained as previously described (21). a-Furfuryl-B-glucoside (2-furanmethanol-B-D-glucopyranoside) was synthesized by the same procedure used for synthesis of FG (21), but using a-furfuryl alcohol as aglycone substrate. All radiochemicals were from the Radiochemical Centre (Amersham, UK), a-and al-furfuryl alcohol were from Aldrich, and silica gel-60 TLC plates (20 x 20) were from Merck. All other chemicals and enzymes were from Sigma. Preparation and Assay of ,-Glucan SynthaseWashed membranes from mung bean hypocotyls were prepared in the presence of PEG-4000 as described (3). Standard assays were performed at 20°C for 10 min and contained in a www.plantphysiol.org on May 12, 2018 -Published by Downloaded from

A membrane-associated form of sucrose synthase and its potential role in synthesis of cellulose and callose in plants.

“…Radioactive derivatives were separated (with similar results) by GLC equipped with a stream splitter (20) or by TLC on silica gel G by using hexane/ethyl acetate, 1:1 (vol/vol), as solvent. Derivatives from unlabeled cellulose, cellobiose, and pachyman were used as standards for terminal Glc and 3-or 4-linked Glc.…”

Characterization of the 1,3- -glucan synthase of Aspergillus fumigatus

“…In contrast to fungal enzymes, plant enzymes require divalent Ca2+ and Mg2+ but do not need GTP for activation. Furfuryl glucoside, which stimulates the glucan synthase of mung bean at least 20-fold (Ohana et al, 1992), has a very low effect (29%) on A . fumigatus glucan synthase (data not shown).…”

Specificity of Binding of β-Glucoside Activators of Ryegrass (1->3)-β-Glucan Synthase and the Synthesis of Some Potential Photoaffinity Activators

“…In the study of Ohana et al (1992) with the cotton (1+3)-p-glucan synthase, ~4-methyl-7-cellobiosylcoumarin ( K , = 40 PM), 4-methyl-7-~-glucosylcoumarin (K, = 45 PM), and P-furfuryl-P-glucoside (K, = 50 PM) were the best activators. In the ryegrass preparation, compounds [15] and [16], both with K,s of 30 piv, were the best activators.…”

“…octyl P-glucoside (in the presence of Ca2+), have mi- 1227 cromolar K,s. P-Furfuryl P-glucoside from Vigna radiata (azuki bean) has a K, of 50 FM with the cotton (Gossypium hirsutum) seed hair (1+3)-P-glucan synthase (Ohana et al, 1991(Ohana et al, , 1992.…”