Cell Wall and Membrane-Associated Exo-β-D-Glucanases from Developing Maize Seedlings

• Published in: Plant physiology (Bethesda) 2000-06, Vol.123 (2), p.471-485
• Main Authors: Jong-Bum Kim, Anna T. Olek, Carpita, Nicholas C.
• Format: Article
• Language: English
• Subjects: Acetates; Amino Acid Sequence; Amino acids; Base Sequence; Biochemical Processes and Macromolecular Structures; Cell Membrane - enzymology; Cell membranes; Cell Wall - enzymology; Cell walls; Cloning, Molecular; Complementary DNA; Corn; DNA Primers; DNA, Complementary; Electrophoresis, Polyacrylamide Gel; Enzymes; Glycoside Hydrolases - genetics; Glycoside Hydrolases - isolation & purification; Glycoside Hydrolases - metabolism; Molecular Sequence Data; Plants; Seedlings; Sequence Homology, Amino Acid; Sodium; Space life sciences; Zea mays - enzymology; Zea mays - growth & development
• ISSN: 0032-0889; 1532-2548
• DOI: 10.1104/pp.123.2.471

A β-D-glucan exohydrolase was purified from the cell walls of developing maize (Zea mays L.) shoots. The cell wall enzyme preferentially hydrolyzes the non-reducing terminal glucosyl residue from (1→3)-β-D-glucans, but also hydrolyzes (1→2)-, (1→6)-, and (1→4)-β-D-glucosyl units in decreasing order of activity. Polyclonal antisera raised against the purified exo-β-D-glucanase (ExGase) were used to select partial-length cDNA clones, and the complete sequence of 622 amino acid residues was deduced from the nucleotide sequences of the cDNA and a full-length genomic clone. Northern gel-blot analysis revealed what appeared to be a single transcript, but three distinct polypeptides were detected in immunogel-blot analyses of the ExGases extracted from growing coleoptiles. Two polypeptides appear in the cell wall, where one polypeptide is constitutive, and the second appears at the time of the maximum rate of elongation and reaches peak activity after elongation has ceased. The appearance of the second polypeptide coincides with the disappearance of the mixed-linkage (1→3),(1→4)-β-D-glucan, whose accumulation is associated with cell elongation in grasses. The third polypeptide of the ExGase is an extrinsic protein associated with the exterior surface of the plasma membrane. Although the activity of the membrane-associated ExGase is highest against (1→3)-β-D-glucans, the activity against (1→4)-β-D-glucan linkages is severely attenuated and, therefore, the enzyme is unlikely to be involved with turnover of the (1→3),(1→4)-β-D-glucan. We propose three potential functions for this novel ExGase at the membrane-wall interface.