Gliadins and glutenins, the major storage proteins of wheat endosperm (Triticum durum, Desf. cv Monroe), were reduced in vitro by the NADP/thioredoxin system (NADPH, NADP-thioredoxin reductase and thioredoxin; in plants, the h type) from either the same source or the bacterium Escherichia coli. A more limited reduction of certain members of these protein groups was achieved with the reduced form of glutathione or glutaredoxin, a protein known to replace thioredoxin in certain bacterial and mammalian enzyme systems but not known to occur in higher plants. Endo The seed is the only tissue for which the NADP/thioredoxin system has been ascribed physiological activity in plants. Thioredoxin h reduces members of several different soluble seed proteins-thionins, a-amylase, and trypsin inhibitors (11, 14)-and also reductively activates an enzyme of carbohydrate metabolism (PPi fructose-6-P, 1-phosphotransferase, or PFP) (13). The results (14) suggest that the inhibitor proteins, long known to be active in bioprotection, may function within the seed to link thioredoxin to the regulation of yet-to-be identified target enzymes (cf. 9, 16, 21).The question arises as to whether thioredoxin can reduce other types of seed proteins. Quantitatively, the most important group is comprised of storage proteins, which account for up to 80% of the total protein of the seed (12,20). In the case of plants such as cereals, these proteins are insoluble in aqueous solutions and are chemically inert until they are reduced. It is not known how these proteins are mobilized during germination, and a physiological agent capable of their reduction has not been described.To help fill this gap, we have undertaken a study with wheat, a cereal with well-characterized seed proteins. We now report that representatives of the major wheat (Triticum durum) storage proteins-the gliadins and glutenins-are specifically reduced by thioredoxin. The results provide evidence that the NADP/thioredoxin system functions in the reduction of the principal seed proteins, thereby increasing their proteolytic susceptibility and making amino acids (nitrogen and sulfur) available during germination. Taken together with our recent work (14), the new findings suggest that thioredoxin functions as a signal to enhance metabolic processes associated with seed germination. A preliminary account of this work has been published (31).
MATERIALS AND METHODS
Plant MaterialSeeds and semolina of durum wheat (Triticum durum, Desf. cv Monroe) were kind gifts of Dr. K. Kahn.
Germination of Wheat SeedsTwenty to thirty seeds were placed in a plastic Petri dish on three layers of Whatman No. 1 filter paper moistened with 5 mL of deionized water. Germination was carried out for up to 4 d at room temperature in a dark chamber. Plant Physiol. Vol. 99, 1992 Reagents/Fine Chemicals Biochemicals and lyophilized coupling enzymes were obtained from Sigma. Escherichia coli thioredoxin and NTR were purchased from American Diagnostica, Inc. (Greenwich, CT). Wheat thioredoxin h and NTR w...
