Purification and Characterization of a Low-Molecular-Weight Xylanase Produced by Acrophialophora nainiana

Abstract

A low-molecular-weight xylanase activity (XynI) was isolated from the fungus Acrophialophora nainiana after growth in a solid medium containing wheat bran. XynI was purified to apparent homogeneity by ultrafiltration and gel filtration chromatography. The purified enzyme had a molecular weight value of approx. 17 kDa, as determined by SDS-PAGE. This enzyme was most active at 50 degreesC and pH 6.0. At 50 degreesC the half-life was 150 min. The apparent Km value for birchwood xylan was much lower than the Km value for oat spelt xylan. XynI was activated by L-cysteine, DTE, beta-mercaptoethanol, and L-tryptophan. XynI did not show significant sequence homology with other xylanases. The analysis of hydrolysis products of xylans and wood pulps showed that XynI was able to release xylooligomers ranging from X2 to X3 and X2 to X6, respectively. The enzyme was not active against acetylated xylan. A small amount of xylose was released from deacetylated, birchwood, and oat spelt xylans. The results obtained with enzymatic treatment of Kraft pulp indicated a reduction in the amount of chlorine compounds required for the process and enhanced brightness gain.

Purification and characterization of a new Xylanase from Humicola grisea var. Thermoidea

“…The purified enzyme gave the highest activity at a temperature range of 55-60ºC. This result was higher than the values reported for some xylanases from Acrophialophora nainiana, Trichoderma harzianum and Penicillium capsulatum (7,18,23). It was lesser than those of xylanase forms 1 and 2 from the same fungus (13) and similar to Xyl1 and Xyl2 from Humicola insolens (6).…”

“…A major xylanase activity was detected in the concentrated crude extract (retentate), while a small amount of enzyme activity was found in the filtrate (permeate). The ability of xylanases to penetrate an ultrafiltration membrane has been reported before (18,19,23). It is suggested to be due to xylanase compact structure and/or non-uniformity of membrane pore size (15,18).…”

“…Ultrafiltration procedure is reported to reduce xylanase recovery yields (15). Low purification yields for xylanase activities are often reported in the literature (7,15,18,22,23).…”

“…However, the hypothesis that the substituents are probably located in regions of the substrate distant from the unsubstituted portions can not be discarded (18). Xyl 1 and Xyl 2 from H. insolens require a sequence of at least three unsubstituted xylose residues for their active binding (23).…”

“…Some properties of these purified enzymes have been published (13,22). Solid-state cultivation on wheat bran as a substrate is an efficient procedure for xylanase production (17,23). Some of the advantages of the use of solidstate cultivation over liquid medium are described such as lower costs and the production of enzymes with higher specific activities (11).…”