Detection of O‐Glycosidically Linked Mannose Within the Structure of a Highly Purified Glutenin Subunit Isolated from Chinese Spring Wheat

Abstract

ABSTRACTA low level of O‐glycosidically linked D‐mannose was detected on the 1Dx2 high Mr glutenin subunit of Chinese Spring wheat. Detection of the O‐glycosidic linkage was accomplished by subjecting the purified 1Dx2 protein to β‐elimination and reduction conditions that resulted in the specific cleavage of O‐glycosidically linked carbohydrates from the protein backbone and the reduction of the linkage (reducing end) sugars to alditols. Subsequently, acid‐catalyzed methanolysis was used to cleave glycosidic linkages on released carbohydrate as well as carbohydrate remaining on the protein. The repeat units were derivatized with trimethylsilyl and analyzed by gas chromatography and mass spectroscopy (GC‐MS). The 1Dx2 high Mr glutenin subunit of Chinese Spring wheat consisted of 1.2% carbohydrate containing D‐mannose and D‐glucose units. O‐Glycosidically linked D‐mannose residues represented 0.13% of the total mass of the 1Dx2 high Mr glutenin subunit. This represented <1 O‐glycosidically linked D‐mannose residue per protein molecule and provided some initial evidence that there may be various glycoforms of this subunit present in the preparation.