Xylan Deconstruction by Thermophilic Thermoanaerobacterium bryantii Hemicellulases Is Stimulated by Two Oxidoreductases

Zhuolin Yi,Abigail E Asangba,Isaac Cann,Dylan Dodd,Siddhartha Chakraborty,Ahmed M Abdel-Hamid,Peter G Stroot,Roderick I Mackie,Xiaoyun Su

doi:10.3390/catal12020182

Abstract

Thermoanaerobacterium bryantii strain mel9T is a thermophilic bacterium isolated from a waste pile of a corn-canning factory. The genome of T. bryantii mel9T was sequenced and a hemicellulase gene cluster was identified. The cluster encodes seven putative enzymes, which are likely an endoxylanase, an α-glucuronidase, two oxidoreductases, two β-xylosidases, and one acetyl xylan esterase. These genes were designated tbxyn10A, tbagu67A, tbheoA, tbheoB, tbxyl52A, tbxyl39A, and tbaxe1A, respectively. Only TbXyn10A released reducing sugars from birchwood xylan, as shown by thin-layer chromatography analysis. The five components of the hemicellulase cluster (TbXyn10A, TbXyl39A, TbXyl52A, TbAgu67A, and TbAxe1A) functioned in synergy to hydrolyze birchwood xylan. Surprisingly, the two putative oxidoreductases increased the enzymatic activities of the gene products from the xylanolytic gene cluster in the presence of NADH and manganese ions. The two oxidoreductases were therefore named Hemicellulase-Enhancing Oxidoreductases (HEOs). All seven enzymes were thermophilic and acted in synergy to degrade xylans at 60 °C. Except for TbXyn10A, the other enzymes encoded by the gene cluster were conserved with high amino acid identities (85–100%) in three other Thermoanaerobacterium species. The conservation of the gene cluster is, therefore, suggestive of an important role of these enzymes in xylan degradation by these bacteria. The mechanism for enhancement of hemicellulose degradation by the HEOs is under investigation. It is anticipated, however, that the discovery of these new actors in hemicellulose deconstruction will have a significant impact on plant cell wall deconstruction in the biofuel industry.

Highlights

On the basis of phylogenetic and physiological differences, we propose that strain mel9T be classified as a new Thermoanaerobacterium species, for which the name Thermoanaerobacterium bryantii is suggested, in honor of the anaerobic microbiologist
The genus Thermoanaerobacterium is, known to be endowed with plant cell wall-degrading enzymes that can be harnessed for enzymatic release of fermentable sugars from plant cell wall polysaccharides, such as cellulose and hemicellulose
Enzymatic deconstruction of plant cell wall polysaccharides is a critical step in the production of second-generation biofuels from

Summary

Introduction

Hemicellulose represents the second most abundant component in plant biomass. Efficient degradation of hemicellulose is critical for economical utilization of the plant cell wall polysaccharides in biofuel production. A major component of hemicellulose is heterogeneous xylan, which contains linear xylose-configured backbones decorated with arabinofuranosyl, glucuronyl, feruloyl, and acetyl group side chains. The complete degradation of heterogeneous xylan into simple sugars requires the synergistic action of a set of hemicellulases [1].

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