PURIFICATION AND CHARACTERIZATION OF A THERMOSTABLE XYLANASE FROM BACILLUS-AMYLOLIQUEFACIENS

A Bacillus amyloliquefaciens strain isolated from soil produced xylano lytic enzymes in the extracellular medium when grown on xylan and nitr ate. No cellulase activity was detected Xylanase was purified from the culture supernatant in three steps which comprised anion-exchange ads orption, ammonium sulfate precipitation and hydrophobic interaction ch romatography. The pure enzyme appeared as two close protein bands oil SDS-PAGE with molecular weights of 18.4 and 19.6 kD, respectively. The molecular weight obtained by sedimentation equilibrium under-native c onditions was about 18.5 kD. The isoelectric point was 10.1. The enzym e contains a relatively high content of aspartate, glycine, and threon ine. Tryptophan seems to be essential for xylanase activity. The prese nce of carbohydrate was noted in the pure enzyme. Circular dichroism s tudies indicated that the protein contains almost equal proportions of alpha-helix, beta-sheet, and beta-turns. The optimum pH of activity w as 6.8-7.0. The enzyme exhibited good stability even at pH 9.0. Excell ent stability was observed at 50 degrees C even though optimal activit y was at 80 degrees C. The activity was completely inhibited by Hg2+ i ons and was reduced drastically in the presence of Cu2+ and Fe3+ ions. Mn2+ ions, EDTA, beta-mercaptoethanol, and dithiothreitol up to 5 mM stimulated the enzyme activity. (C) 1998 Elsevier Science Inc.