Polypeptide characteristics and immunological properties of exo- and endoglucanases purified from maize coleoptile cell walls.

摘要

Cereal coleoptile cell walls have exo- and endoglucanases [cellulases] capable of mediating the hydrolysis of non-cellulosic beta-(1,3)(1,4)-glucan in situ. A purified exoglucanase (EC 3.2.1.58) resolved as a single band at 73.5 kDa, while endoglucanase isoenzymes consistently appeared as two bands at 32.9 and 34.3 kDa when subjected to SDS-PAGE. HPLC analysis of the native proteins by gel-permeation chromatography revealed molecular weights of about 55 and 29 kDa for the exo- and endoglucanases, respectively. The exoglucanase has an isoelectric focusing point at pI 7.2 and the endoglucanase isoenzymes appeared as two major bands, one at pI 7.8 and another at 7.3. Deglycosylation of the native proteins followed by SDS-PAGE demonstrated that sugarsaccounted for about 6.5% of the exoglucanase and were 12.5 and 8.8% of the two endoglucanase isoenzymes, respectively. After deglycosylation the two endoglucanases converged at 30.0 kDa, suggesting polypeptide homology and that divergence in electrophoretic mobility was a consequence of glycosylation. Antibodies raised against intact exo- and endoglucanases recognized the polypeptide of the corresponding enzymes, irrespective of glycosylation. The N-terminal amino acid sequence supported the conclusionthat the exo- and endoglucanase have different polypeptide structures.