α-Mannosidase (EC 3.2.1.24) from rice dry seeds was purified to homogeneity. Optimum pH and Km for pNP-α-Man hydrolysis were pH 4.3-4.5 and 1.04 mM, respectively. The enzyme digested mannobioses such as Manα-1,2Man, Manα1,6Man, Manα-1,3Man but Manα-1,4Man. Zn2+ ion was required for the activity, whereas EDTA and swainsonine inhibited the activity by 80 and 96%, respectively. The rice storage protein, glutelin was prepared and its basic subunits were shown to have high mannose-type sugar chains by two-dimensional mapping using NH2-P and C18 silica columns. They were Man9GlcNAc2, Man8GlcNAc2, Man7GlcNAc2, Man6GlcNAc2 and Man5GlcNAc2. All these oligosaccharides were digested by the purified α-mannosidase, and ManGlcNAc2 and mannose were formed. Glycopeptides, having these high mannose-type sugar chains, could also be digested by the α-mannosidase. Subunits were prepared from glutelin basic subunit and the richest subunit among them, subunit 2 (isoform 2), was digested by the α-mannosidase. Isoform 2 was digested by V8 protease only partially and slowly. However, isoform 2, pre-treated with the α-mannosidase, was rapidly and completely digested by V8 protease.