Functional characterization of a novel GH94 glycoside phosphorylase, 3-O-β-D-glucopyranosyl β-D-glucuronide phosphorylase, and implication of the metabolic pathway of acidic carbohydrates in Paenibacillus borealis

Glycoside hydrolase family 94 (GH94) contains enzymes that reversibly catalyze the phosphorolysis of beta-glycosides. We conducted this study to investigate a GH94 protein (PBOR_13355) encoded in the genome of Paenibacillus borealis DSM 13188 with low sequence identity to known phosphorylases. Screening of acceptor substrates for reverse phosphorolysis in the presence of alpha-D-glucose 1-phosphate as a donor substrate showed that PBOR_13355 utilized D-glucuronic acid and p-nitrophenyl beta-D-glucuronide as acceptors. In the reaction with D-glucuronic acid, 3-O-beta-D-glucopyranosyl-D-glucuronic acid was synthesized. PBOR_13355 showed a higher apparent catalytic efficiency to p-nitrophenyl beta-D-glucuronide than to D-glucuronic acid, and thus, PBOR_13355 was concluded to be a novel glycoside phosphorylase, 3-O-beta-D-glucopyranosyl beta-D-glucuronide phosphorylase. PBOR_13360, encoded by the gene immediately downstream of the PBOR_13355 gene, was shown to be beta-glucuronidase. Collectively, PBOR_13355 and PBOR_13360 are predicted to work together in the cytosol to metabolize oligosaccharides containing the 3-O-beta-D-glucopyranosyl beta-D-glucuronide structure released from bacterial and plant acidic carbohydrates. (C) 2022 Elsevier Inc. All rights reserved.

Automated summary

A novel glycoside phosphorylase PBOR_13355 and beta-glucuronidase PBOR_13360 were identified in this study. They are predicted to work together in the cytosol to metabolize acidic carbohydrates.