GH28 polygalacturonase of human gut Bacteroides spp. confers the degradation of a novel branched galacturonan isolated from Jasminum nudiflorum.

The specific enzymes responsible for metabolizing distinct pectin structures have not been fully elucidated due to the structural diversity of pectin. Thus, we hypothesize that there may be other enzymes to target pectin. Here, a novel pectic polysaccharide CFM224 (18.2 kDa) was isolated from Jasminum nudiflorum for the first time. Structural analysis revealed a unique structure comprising branched galacturonan backbone interspersed with 2-α-Rhap, while 2, 4-α-Rhap existed on the branch. Complex branches including arabinogalactan, glucan polymers and t-GalpA, ΔHexpA, the other was composed of 4-β-Glcp, 2, 4-α-Rhap, t-β-Rhap and t-GalpA. CFM224 promoted the growth of Bacteroides spp. and underwent degradation. Furthermore, two polygalacturonases belonging to the glycoside hydrolase 28 (GH28) family, which degraded CFM224 into GalA, encoded by BXY_32940 and BT_4123 genes of Bacteroides spp., respectively, were identified. Notably, the predicted structures of BXY_32940GH28 and BT_4123GH28 exhibit significant homology to rhamnogalacturonase. Site mutation and enzymatic activity analysis verified that D312 might be the catalytic residue of GH28. Genetic analysis demonstrated that polygalacturonase was predominantly encoded by Bacteroides spp. Overall, these findings further expand the metabolic network of pectin and may provide novel candidates for prebiotic screening.