Intracellular debranching enzymes from Cellulomonas sp. B6 enable efficient arabino- and glucuronoxylan breakdown

Abstract

Xylan constitutes a major portion of plant biomass, typically accounting for 20% to 35% of the dry weight in hardwoods and grasses. The sustainable valorization of lignocellulosic biomass requires efficient xylan deconstruction that goes beyond the action of canonical xylanases. Here, two intracellular glycoside hydrolases (GHs) from Cellulomonas sp. B6 were characterized: a GH51 subfamily 1 a-L-arabinofuranosidase (CsAbf51A) and a GH67 a-D-glucuronidase (CsAgu67) that target key glycosyl substitutions of xylan. CsAbf51A exhibited exo-activity on a(1,2), a(1,3), and a(1,5) arabinose residues from mono-substituted arabino-xylooligosaccharides (AXOS) and arabinans, showing a preference for these substrates over polymeric arabinoxylan. CsAgu67 functioned as a strict exo-a-1,2-glucuronidase, releasing 4-O-methyl-a-D-glucuronic acid (MeGlcA) exclusively from the non-reducing ends of glucurono-xylooligosaccharides (UXOS) but not from polymeric glucuronoxylan. Such substrate selectivity aligns with a metabolic strategy that involves the internalization of substituted XOS followed by further processing inside the cell. When incorporated into xylanolytic cocktails with CsXyn10A (a GH10 xylanase) and EcXyl43 (a GH43 b-xylosidase), xylose release was synergically boosted from diverse glucurono- and arabinoxylan-rich biomasses. This study demonstrates that these intracellular debranching enzymes, often overlooked in bacterial metabolism, can be used to expand the biocatalytic repertoire for xylan valorization. Furthermore, their integration into tailored enzyme formulations provides a powerful strategy to advance the development of efficient biorefineries.