Purification and characterization of xylanases from Cellulomonas sp. N.C.I.M. 2353

Cellulomonas produces multiple xylanases in the presence of xylan, carboxymethylcellulose (CMC), starch and, to a very much lesser extent, cellobiose. A procedure involving (NH₄)₂SO₄ precipitation, DEAE-cellulose chromatography and gel filtration was used to purify the xylanases. True xylanases and xylanases with multiple specificity were detected. True xylanases had an optimum pH of 6.5, a temperature optimum of about 55°C, and pI values of about 8.0. The molecular masses of the purified xylanases were found to be 23, 33 and 53 kDa. The K(m) and V(max) values of the 33 and 53 kDa enzymes were 1.7 and 1.5 mg/ml, and 380 and 690 μmol/min per mg respectively. The 33 and 53 kDa enzymes were totally inactivated in the presence of HgCl₂ and AgNO₃. These two enzymes were not affected by 5 mM concentrations of KCl, MgCl₂, NaCl and MnSO₄. These enzymes were identified as endoxylanases on the basis of their end products. Xylanases other than these three enzymes were also found in a 'cellulosome' type of complex in Cellulomonas. Enzymes in this complex showed activity towards both CMC and xylan.