Purification and characterization of two thermostable xylanases from Paenibacillus sp. DG-22

Two thermostable xylanases, designated XynA and XynB, were purified to homogeneity from the culture supernatant of Paenibacillus sp. DG-22 by ion-exchange and gel-filtration chromatography. The molecular masses of xylanases A and B were 20 and 30 kDa, respectively, as determined by SDS-PAGE, and their isoelectric points were 9.1 and 8.9, respectively. Both enzymes had similar pH and temperature optima (pH 5.0- 6.5 and 70°C), but their stability at various temperatures differed. Xylanase B was comparatively more stable than xylanase A at higher temperatures. Xylanases A and B differed in their K_m and V_max values. XynA had a K_m of 2.0 mg/ml and a V_max of 2,553 U/mg, whereas XynB had a K_m of 1.2 mg/ml and a V_max of 754 U/mg. Both enzymes were endo-acting, as revealed by their hydrolysis product profiles on birchwood xylan, but showed different modes of action. Xylotriose was the major product of XynA activity, whereas XynB produced mainly xylobiose. These enzymes utilized small oligosaccharides such as xylotriose and xylotetraose as substrates, but did not hydrolyzed xylobiose. The amino terminal sequences of XynA and XynB were determined. Xylanase A showed high similarity with low molecular mass xylanases of family 11.