Abstract
BackgroundXylanases randomly cleave the internal β-1,4-glycosidic bonds in the xylan backbone and are grouped into different families in the carbohydrate-active enzyme (CAZy) database. Although multiple xylanases are detected in single strains of many filamentous fungi, no study has been reported on the composition, synergistic effect, and mode of action in a complete set of xylanases secreted by the same microorganism.ResultsAll three xylanases secreted by Penicillium chrysogenum P33 were expressed and characterized. The enzymes Xyl1 and Xyl3 belong to the GH10 family and Xyl3 contains a CBM1 domain at its C-terminal, whereas Xyl2 belongs to the GH11 family. The optimal temperature/pH values were 35 °C/6.0, 50 °C/5.0 and 55 °C/6.0 for Xyl1, Xyl2, and Xyl3, respectively. The three xylanases exhibited synergistic effects, with the maximum synergy observed between Xyl3 and Xyl2, which are from different families. The synergy between xylanases could also improve the hydrolysis of cellulase (C), with the maximum amount of reducing sugars (5.68 mg/mL) observed using the combination of C + Xyl2 + Xyl3. Although the enzymatic activity of Xyl1 toward xylan was low, it was shown to be capable of hydrolyzing xylooligosaccharides into xylose. Xyl2 was shown to hydrolyze xylan to long-chain xylooligosaccharides, whereas Xyl3 hydrolyzed xylan to xylooligosaccharides with a lower degree of polymerization.ConclusionsSynergistic effect exists among different xylanases, and it was higher between xylanases from different families. The cooperation of hydrolysis modes comprised the primary mechanism for the observed synergy between different xylanases. This study demonstrated, for the first time, that the hydrolysates of GH11 xylanases can be further hydrolyzed by GH10 xylanases, but not vice versa.
Highlights
Xylanases randomly cleave the internal β-1,4-glycosidic bonds in the xylan backbone and are grouped into different families in the carbohydrate-active enzyme (CAZy) database
In the phylogenetic tree reconstructed using the neighbor-joining method (Fig. 1), Xyl1 and Xyl3 were grouped into the GH10 family, whereas Xyl2 was grouped into the GH11 family according to the domain
Both Xyl1 and Xyl3 are GH10 xylanases, Xyl3 possesses an extra loop compared with Xyl1 (Fig. 2)
Summary
Xylanases randomly cleave the internal β-1,4-glycosidic bonds in the xylan backbone and are grouped into different families in the carbohydrate-active enzyme (CAZy) database. Yang et al Microb Cell Fact (2019) 18:159 exhibit low substrate specificity and have high affinity toward the highly branched xylan backbone [6]. These enzymes have small substrate binding sites that have a high affinity toward shorter linear xylooligosaccharides [7, 8]. Multiple xylanases with different cleavage patterns of xylosidic bonds are required to increase the extent of xylan hydrolysis to monosaccharide [15]. For this reason, many filamentous fungi may produce different xylanases in the presence of lignocellulosic materials. An understanding of this process may reveal the mechanism of hemicellulose degradation by microorganisms and show their adaptability to the natural environment
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