Molecular details of a starch utilization pathway in the human gut symbiont Eubacterium rectale

Abstract

Summary Eubacterium rectale is a prominent human gut symbiont yet little is known about the molecular strategies this bacterium has developed to acquire nutrients within the competitive gut ecosystem. Starch is one of the most abundant glycans in the human diet, and E. rectale increases in vivo when the host consumes a diet rich in resistant starch, although it is not a primary degrader of this glycan. Here we present the results of a quantitative proteomics study in which we identify two glycoside hydrolase 13 family enzymes, and three ABC transporter solute-binding proteins that are abundant during growth on starch and, we hypothesize, work together at the cell surface to degrade starch and capture the released maltooligosaccharides. EUR_21100 is a multidomain cell wall anchored amylase that preferentially targets starch polysaccharides, liberating maltotetraose, while the membrane associated maltogenic amylase EUR_01860 breaks down maltooligosaccharides longer than maltotriose. The three solute-binding proteins display a range of glycan-binding specificities that ensure the capture of glucose through maltoheptaose and some α1,6-branched glycans. Taken together, we describe a pathway for starch utilization by E. rectale DSM 17629 that may be conserved among other starch-degrading Clostridium cluster XIVa organisms in the human gut.

Novel carbohydrate binding modules in the surface anchored α‐amylase of Eubacterium rectale provide a molecular rationale for the range of starches used by this organism in the human gut

“…We previously reported that E. rectale Amy13K (EUR_21100) was likely comprised of five CBMs at its N‐terminus based on weak sequence homology to the starch‐binding families CBM26 (BLAST E ‐value 2e −5 ) and CBM41 (BLAST E ‐value 1e −5 – 5e −7 ) (Cockburn et al , ). The biochemical and structural data presented here supports that there are five CBMs, labeled as CBMa‐e and two warrant classification into new CBM families (Fig.…”

“…For activity assays with polysaccharide substrates the production of free reducing ends was monitored using the bicinchoninic acid (BCA) method (Waffenschmidt and Jaenicke, ) as previously described (Cockburn et al , ). All reactions included 10 mM HEPES pH 6.5, with 5 mM CaCl 2 and 0.02% Tween80.…”

An Extracellular Cell-Attached Pullulanase Confers Branched α-Glucan Utilization in Human Gut Lactobacillus acidophilus

“…Ruminococcus bromii has been identified as the primary degrader of resistant starch in the human gut (23), although Bifidobacterium adolescentis strains were also reported to possess growth capabilities on this substrate (15). Major commensals from the Bacteroides genus (47) and the butyrate-producing Firmicutes member Eubacterium rectale (14) are other HGM members with considerable starch growth and degradation capabilities. Common to these bacteria is that they possess highly modular extracellular cell-attached enzymes with one or more catalytic modules and multiple carbohydrate binding modules (CBMs), which mediate tight binding to starch substrates.…”

The Sus operon: a model system for starch uptake by the human gut Bacteroidetes

“…In addition to the diversity of enzymes complements employed, the strategies used to capture hydrolyzed starch in the gut are a function of the unique physiology of the respective microorganisms [14–17]. The Gram-positive Firmicutes and Actinobacteria take up monosaccharides and oligosaccharides via a variety of transport systems including ATP-binding cassette transporters, major facilitator superfamily, and phosphotransferase systems [18, 19].…”

“…The Gram-positive Firmicutes and Actinobacteria take up monosaccharides and oligosaccharides via a variety of transport systems including ATP-binding cassette transporters, major facilitator superfamily, and phosphotransferase systems [18, 19]. Many of these transporters are encoded within putative operons that include one or more extracellular GH13 enzymes to hydrolyze starch at the cell surface [14, 17, 20]. In contrast, the genomes of most Bacteroidetes, the dominant Gram-negative phylum in the mammalian gut, have far fewer of these classically studied carbohydrateuptake systems [21].…”