Plant N-glycan breakdown by human gut Bacteroides

Abstract

The major nutrients available to the human colonic microbiota are complex glycans derived from the diet. To degrade this highly variable mix of sugar structures, gut microbes have acquired a huge array of different carbohydrate-active enzymes (CAZymes), predominantly glycoside hydrolases, many of which have specificities that can be exploited for a range of different applications. Plant N-glycans are prevalent on proteins produced by plants and thus components of the diet, but the breakdown of these complex molecules by the gut microbiota has not been explored. Plant N-glycans are also well characterised allergens in pollen and some plant-based foods, and when plants are used in heterologous protein production for medical applications, the N-glycans present can pose a risk to therapeutic function and stability. Here we use a novel genome association approach for enzyme discovery to identify a breakdown pathway for plant complex N-glycans encoded by a gut Bacteroides species and biochemically characterise five CAZymes involved, including structures of the PNGase and GH92 α-mannosidase. These enzymes provide a toolbox for the modification of plant N-glycans for a range of potential applications. Furthermore, the keystone PNGase also has activity against insect-type N-glycans, which we discuss from the perspective of insects as a nutrient source.

N-glycan breakdown by bacterial CAZymes

“…PNGaseF-II ( E. meningoseptica ) also has specificity towards N-glycans with either α1,3- and α1,6-fucose on the core GlcNAc, but has a strong preference for denatured protein [ 10 ]. B035DRAFT_03341 PNGase from gut commensal Bacteroides massiliensis is selective for N-glycans with α1,3-fucose, and BF0811 PNGase from Bacteroides fragilis has very similar activity to PNGaseF [ 11 ].…”

“…For instance, three GH92 enzymes from B. thetaiotaomicron that are in the same operon as the GH18 BT3987 act in sequence to remove the α1,2-, α1,3-, and the first α1,6-mannose to leave Man2 ( Figure 3 ) [ 12 ]. In terms of plant N-glycans, the human gut commensal B. massiliensis and the plant pathogen Xanthomonas campestris have both been shown to encode a GH92 with specificity towards the core α1,3-mannose in plant N-glycans, so it is able to accommodate the β1,2-xylose in its binding site [ 11 ]. A recent study of three GH38 enzymes and a GH125 enzyme from Bifidobacterium longum , whose genes are encoded in the same operon, shows overlapping activities towards the linkages in high-mannose N-glycans ( Figure 3 ) [ 29 ].…”

“…Hydrolysis of core α1,3-fucose decorations characteristic of plant and invertebrate N-glycans has also been observed in GH29 enzymes from the human gut commensal B. massiliensis and plant pathogen X. campestris [ 11 , 45 ]. These enzymes require the N-glycan to be significantly processed before acting, suggesting that there is a strict sequence of deconstruction by these organisms for this substrate.…”

“…These enzymes require the N-glycan to be significantly processed before acting, suggesting that there is a strict sequence of deconstruction by these organisms for this substrate. In terms of the plant complex N-glycan α1,4-linked antennary fucose, two GH29 enzymes were identified from B. massiliensis that can remove this decoration from the full plant N-glycan structure [ 11 ].…”

“…( C ) Useful glycan structures discussed in the review. ( D ) Examples of glycan structures adapted from [ 11 ] from a variety of organisms displaying nonclassical compositions [ 54–59 ].…”