TERTIARY STRUCTURE OF XYLANASE AND ESTIMATION OF ACTIVE SITES BY SITE DIRECTED MUTAGENESIS

The nucleotide sequence of xylanase gene(xynA) of Bacillus pumilus IPO, a hyperproducer of xylanase, was determined and the amino acid sequence was deduced from it. Xylanase is produced as a preenzyme consisting of the mature enzyme of 201 amino acid residues and a signal peptide of 27 residues. Xylanase was analyzed by X-ray crystallography at the level of 2.2 A resolution. It is consisted with two domains, smaller and larger, between two a crevasse suitable to accept xylan molecule was observed. The mutant xylanases obtained by site directed mutagenesis having amino acid alteration; Glu93-Ser93 and Glul82_Asp182 had no catalytic activity (less than 1/10,000)