Purification of Xyloglucan Hydrolase/Endotransferase from Cell Walls of Azuki Bean Epicotyls

An enzyme involved in the breakdown of xyloglucans was purified from an extract of cell walls of azuki bean epicotyls obtained with 1 M NaCl and purified by column chromatography on several different resins. The purified enzyme gave a single band of a protein with a molecular mass of about 32 kDa after SDS-PAGE. The enzyme hydrolyzed the xyloglucans of high molecular mass from azuki cell walls to yield fragments of about 50 kDa without production of any oligo- or monosaccharides. Moreover, the enzyme had hardly any effect on xyloglucans of less than 60 kDa. The enzyme also hydrolyzed xyloglucans from tamarind, but it did not react with cellulose derivatives. In the presence of pyridylamino-labeled xyloglucan oligosac-charides as acceptor substrates, the enzyme catalyzed the transfer of 50-kDa products to the oligosaccharides. The Km value of the enzyme for xyloglucans of 540 kDa was similar in the presence and in the absence of xyloglucan oligosaccharides as acceptors: 1.0 mg ml−1. These results suggest that the enzyme was an endotransferase but had unusual acceptor specificity, preferring smaller acceptors such as water.