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Characterization of XYN10B, a modular xylanase from the ruminal protozoan Polyplastron multivesiculatum, with a family 22 carbohydrate-binding module that binds to cellulose.

ABSTRACT: A new xylanase gene, xyn10B, was isolated from the ruminal protozoan Polyplastron multivesiculatum and the gene product was characterized. XYN10B is the first protozoan family 10 glycoside hydrolase characterized so far and is a modular enzyme comprising a family 22 carbohydrate-binding module (CBM) preceding the catalytic domain. The CBM22 was shown to be a true CBM. It showed high affinity for soluble arabinoxylan and is the first example of a CBM22 that binds strongly to celluloses of various crystallinities. The enzymic properties of XYN10B were also analysed. Its optimal temperature and pH for activity were 39 degrees C and 7.0 respectively; these values being close to those of the ruminal ecosystem. The phylogenetic relationships between the XYN10B CBM22 or catalytic domain and related sequences from ruminal and non-ruminal bacteria and eukaryotes are reported. The xyn10B gene is shown to lack introns.

SUBMITTER: Devillard E

PROVIDER: S-EPMC1223500 | biostudies-other | 2003 Jul

REPOSITORIES: biostudies-other

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Project description:BackgroundEnzymatic removal of hemicellulose components such as xylan is an important factor for maintaining high glucose conversion from lignocelluloses subjected to low-severity pretreatment. Supplementation of xylanase in the cellulase mixture enhances glucose release from pretreated lignocellulose. Filamentous fungi produce multiple xylanases in their cellulase system, and some of them have modular structures consisting of a catalytic domain and a family 1 carbohydrate-binding module (CBM1). However, the role of CBM1 in xylanase in the synergistic hydrolysis of lignocellulose has not been investigated in depth.ResultsThermostable endo-β-1,4-xylanase (Xyl10A) from Talaromyces cellulolyticus, which is recognized as one of the core enzymes in the fungal cellulase system, has a modular structure consisting of a glycoside hydrolase family 10 catalytic domain and CBM1 at the C-terminus separated by a linker region. Three recombinant Xyl10A variants, that is, intact Xyl10A (Xyl10Awt), CBM1-deleted Xyl10A (Xyl10AdC), and CBM1 and linker region-deleted Xyl10A (Xyl10AdLC), were constructed and overexpressed in T. cellulolyticus. Cellulose-binding ability of Xyl10A CBM1 was demonstrated using quartz crystal microbalance with dissipation monitoring. Xyl10AdC and Xyl10AdLC showed relatively high catalytic activities for soluble and insoluble xylan substrates, whereas Xyl10Awt was more effective in xylan hydrolysis of wet disc-mill treated rice straw (WDM-RS). The enzyme mixture of cellulase monocomponents and intact or mutant Xyl10A enhanced the hydrolysis of WDM-RS glucan, with the most efficient synergism found in the interactions with Xyl10Awt. The increased glucan hydrolysis yield exhibited a linear relationship with the xylan hydrolysis yield by each enzyme. This relationship revealed significant hydrolysis of WDM-RS glucan with lower supplementation of Xyl10Awt.ConclusionsOur results suggest that Xyl10A CBM1 has the following two roles in synergistic hydrolysis of lignocellulose by Xyl10A and cellulases: enhancement of lignocellulosic xylan hydrolysis by binding to cellulose, and the efficient removal of xylan obstacles that interrupt the cellulase activity (because of similar binding target of CBM1). The combination of CBM-containing cellulases and xylanases in a fugal cellulase system could contribute to reduction of the enzyme loading in the hydrolysis of pretreated lignocelluloses.

Dataset Information

Characterization of XYN10B, a modular xylanase from the ruminal protozoan Polyplastron multivesiculatum, with a family 22 carbohydrate-binding module that binds to cellulose.

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