Xylanases excreted by the fungus Sporotrichum dimorphosporum have been separated by chromatography on DEAE-Sephadex at pH 7 into Fractions I and II. Gel filtration did not further resolve the enzymes, and preparative electrofocusing was necessary to isolate xylanases of p1 7.1, 7.2, 7.35, and 7.95 in fraction I and p1 3.9, 4,4.4, 4.7, and 5.5 in fraction II. The latter fraction was studied in detail, and afforded two pure xylanases (~14.4 and 4.7); the others were contaminated by O-(carboxymethyl)cellulases.
The xylanases of p1 3.9, 4, 4.4, and 4.7 had molecular weights of 32,000 and that of the xylanase of pI 5.5 was 26,000. The optimal reaction temperature was 65-70" for the former enzymes and 50" for the latter. The optimum pH of action was 4.5-5 for all of these xylanases. The xylanases of ~14.7 and 5.5, the major constituents of the system, exhibit different properties and different values of their Michaelis constants.
The end-products of the reaction affect the reactivity of the enzymes. Xylobiose and 4-thioxylobiose are competitive inhibitors of the xylanase of ~15.5, but they are activators of the xylanase of ~14.7 in the presence of xylan. This observation is suggestive of a new mechanism for regulation of hydrolysis.