α-Mannosidase from the seeds of Triticale

Abstract

Seeds of Triticale (hybrid of wheat and rye) contain an N-acetylglucosamine specific lectin that was affinity purified in our laboratory (Siva Kumar, N. and Padma, K. (1996) "Affinity purification of N-acetyl glucosamine specific lectin. Purification and partial characterization of Triticale lectin". Biochem. Mol. Biol. Int. 38, 1059-1066). Seed extracts also exhibited α-mannosidase activity that was isolated by a combination of ion exchange, hydrophobic chromatography and gel filtration. The purified enzyme is a glycoprotein with 7% carbohydrate and exhibited a native molecular mass of 1,95,000 (±5000) on Biogel P-200 and dissociated into two major subunits under reducing conditions of molecular masses 58 and 40 kDa, respectively. Both subunits cross-reacted with an antibody to the well-characterized jack bean α-mannosidase, suggesting antigenic similarity between the legume and the cereal mannosidases. Purified enzyme binds to Con A-Sepharose gel, possibly through the sugar-binding site. Purified Triticale enzyme was stable at 50°C up to 20 min and did not show requirement of metal ions for activity. Phenylalanine was detected as the sole N-terminal amino acid in the purified enzyme.