STRUCTURE OF THE XYLANASE FROM PENICILLIUM-SIMPLICISSIMUM

Despite its relatively low pH and temperature optimum, the xylanase fr om Penicillium simplicissimum performs exceedingly well under conditio ns of paper bleaching. We have purified and characterized this enzyme, which belongs to family 10 of glycosyl hydrolases. Its gene was clone d, and the sequence of the protein was deduced from the nucleotide seq uence. The xylanase was crystallized from ammonium sulfate at pH 8.4, and X-ray data were collected at cryo-temperature to a crystallographi c resolution of 1.75 Angstrom. The crystal structure was solved by mol ecular replacement using the catalytic domain of the Clostridium therm ocellum xylanase as a search model, and refined to a residual of R = 2 0% (R-free = 23%) for data between 10 and 1.75 Angstrom. The xylanase folds in an (alpha/beta)(8) barrel (TIM-barrel), with additional helic es and loops arranged at the ''top'' forming the active site cleft. In its overall shape, the P. simplicissimum xylanase structure is simila r to other family 10 xylanases, but its active site cleft is much shal lower and wider. This probably accounts for the differences in catalys is and in the mode of action of this enzyme. Three glycerol molecules were observed to bind within the active site groove, one of which inte racts directly with the catalytic glutamate residues. It appears that they occupy putative xylose binding subsites.