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Rice α ‐mannosidase digesting the high mannose glycopeptide of glutelin
Author(s) -
Kishimoto Tadashi,
Hori Hidetaka,
Takano Daisuke,
Nakano Yoshiyasu,
Watanabe Mayumi,
Mitsui Toshiaki
Publication year - 2001
Publication title -
physiologia plantarum
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.351
H-Index - 146
eISSN - 1399-3054
pISSN - 0031-9317
DOI - 10.1034/j.1399-3054.2001.1120103.x
Subject(s) - glutelin , mannosidase , mannose , protease , chemistry , protein subunit , biochemistry , glycopeptide , hydrolysis , oligosaccharide , enzyme , storage protein , chromatography , gene , antibiotics
α ‐Mannosidase (EC 3.2.1.24) from rice dry seeds was purified to homogeneity. Optimum pH and K m for pNP‐ α ‐Man hydrolysis were pH 4.3–4.5 and 1.04 m M , respectively. The enzyme digested mannobioses such as Man α ‐1,2Man, Man α ‐1,6Man, Man α ‐1,3Man but Man α ‐1,4Man. Zn 2+ 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 NH 2 ‐P and C 18 silica columns. They were Man 9 GlcNAc 2 , Man 8 GlcNAc 2 , Man 7 GlcNAc 2 , Man 6 GlcNAc 2 and Man 5 GlcNAc 2 . All these oligosaccharides were digested by the purified α ‐mannosidase, and Man‐GlcNAc 2 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.