{"paper_id":"123833a8-c65c-4dc8-b949-aa69972a1aae","body_text":"Abstract\nβ-Mannans are widespread in the human diet as components of plant-derived foods and as food additives. Several classes of β-mannans are decorated with acetylations, which are key for their physicochemical properties and protection against enzymatic degradation. While the process for depolymerization of acetylated β-mannans has been described in depth for members of the phylum Bacillota, there is limited mechanistic knowledge on how Bacteroidota utilize these glycans. Here, we combined proteomics and biochemical analyses to functionally characterize a pair of carbohydrate esterases (CEs) from Bacteroides cellulosilyticus that, together, deacetylate complex β-mannans. We demonstrate that the newly identified BcCExxx enzyme, representing a novel carbohydrate esterase (CE) family, exhibits high specificity by selectively removing axially oriented 2-O-acetyl groups from mannose residues. In contrast, BcCE7 functions as a broad-spectrum esterase, capable of deacetylating oligosaccharides derived from structurally diverse substrates, including β-mannans, xylans, and acetylated cellulose. In transesterification reactions, BcCExxx showed activity on both mannooligosaccharides and polymeric glucomannan. Overall, our findings provide new insight into the strategies that beneficial Bacteroides have evolved to deacetylate complex β-mannans in the human gut.\nSignificance Statement β-Mannans, commonly found in various plant-based foods and used as food additives, can be metabolized by gut microbes, potentially impacting host health. Acetylation of β-mannans enhances their resistance to enzymatic breakdown, making carbohydrate esterases crucial for gut bacteria to utilize these carbohydrates. This study explores the roles of two carbohydrate esterases (CEs) from a human commensal Bacteroides species in deacetylating complex β-mannans. The two esterases together remove acetylations at different positions on mannose units. BcCE7 is a versatile esterase that deacetylates a variety of oligosaccharides, while the newly discovered BcCExxx is highly active towards 2-O-acetylations on mannose units in both oligosaccharides and polymeric β-mannan. This research enhances our understanding of how Bacteroidota species deacetylate β-mannans.\nCompeting Interest Statement\nThe authors have declared no competing interest.\nFootnotes\nCompeting Interest Statement: The authors declare no competing interests.\nClassification: Biological Sciences/Biochemistry.","source_license":"CC-BY-4.0","license_restricted":false}