The human gut microbe Bacteroides thetaiotaomicron encodes the founding member of a novel glycosaminoglycan-degrading polysaccharide lyase family PL29

Glycosaminoglycans (GAGs) and GAG-degrading enzymes have wide-ranging applications in the medical and biotechnological industries. The former are also an important nutrient source for select species of the human gut microbiota (HGM), a key player in host–microbial interactions. How GAGs are metabolized by the HGM is therefore of interest and has been extensively investigated in the model human gut microbe Bacteroides thetaiotaomicron. The presence of as-yet uncharacterized GAG-inducible genes in its genome and of related species, however, is testament to our incomplete understanding of this process. Nevertheless, it presents a potential opportunity for the discovery of additional GAG-degrading enzymes. Here, we investigated a gene of unknown function (BT_3328) from the chondroitin sulfate (CS) utilization locus of B. thetaiotaomicron. NMR and UV spectroscopic assays revealed that it encodes a novel polysaccharide lyase (PL), hereafter referred to as BtCDH, reflecting its source (B. thetaiotaomicron (Bt)) and its ability to degrade the GAGs CS, dermatan sulfate (DS), and hyaluronic acid (HA). When incubated with HA, BtCDH generated a series of unsaturated HA sugars, including Δ4,5UA-GlcNAc, Δ4,5UA-GlcNAc-GlcA-GlcNac, Δ4,5UA-[GlcNAc-GlcA]2-GlcNac, and Δ4,5UA-[GlcNAc-GlcA]3-GlcNac, as end products and hence was classed as endo-acting. A combination of genetic and biochemical assays revealed that BtCDH localizes to the cell surface of B. thetaiotaomicron where it enables extracellular GAG degradation. BtCDH homologs were also detected in several other HGM species, and we therefore propose that it represents the founding member of a new polysaccharide lyase family (PL29). The current discovery also contributes new insights into CS metabolism by the HGM.

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Structural basis for the exolytic activity of polysaccharide lyase family 6 alginate lyase BcAlyPL6 from human gut microbe Bacteroides clarus

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2021.02.040 ◽

2021 ◽

Vol 547 ◽

pp. 111-117

Author(s):

Bing Wang ◽

Sheng Dong ◽

Fu-Li Li ◽

Xiao-Qing Ma

Keyword(s):

Alginate Lyase ◽

Structural Basis ◽

Human Gut ◽

Polysaccharide Lyase ◽

Gut Microbe

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Ascertaining the biochemical function of an essential pectin methylesterase in the gut microbe Bacteroides thetaiotaomicron

Journal of Biological Chemistry ◽

10.1074/jbc.ra120.014974 ◽

2020 ◽

Vol 295 (52) ◽

pp. 18625-18637

Author(s):

Cheng-Jie Duan ◽

Arnaud Baslé ◽

Marcelo Visona Liberato ◽

Joseph Gray ◽

Sergey A. Nepogodiev ◽

...

Keyword(s):

Gut Microbiota ◽

Critical Role ◽

Pectin Methylesterase ◽

Bacteroides Thetaiotaomicron ◽

Human Gut ◽

Human Gut Microbiota ◽

New Family ◽

Gut Microbe ◽

Dietary Nutrient ◽

Helical Proteins

Pectins are a major dietary nutrient source for the human gut microbiota. The prominent gut microbe Bacteroides thetaiotaomicron was recently shown to encode the founding member (BT1017) of a new family of pectin methylesterases essential for the metabolism of the complex pectin rhamnogalacturonan-II (RG-II). However, biochemical and structural knowledge of this family is lacking. Here, we showed that BT1017 is critical for the metabolism of an RG-II–derived oligosaccharide ΔBT1017oligoB generated by a BT1017 deletion mutant (ΔBT1017) during growth on carbohydrate extract from apple juice. Structural analyses of ΔBT1017oligoB using a combination of enzymatic, mass spectrometric, and NMR approaches revealed that it is a bimethylated nonaoligosaccharide (GlcA-β1,4-(2-O-Me-Xyl-α1,3)-Fuc-α1,4-(GalA-β1,3)-Rha-α1,3-Api-β1,2-(Araf-α1,3)-(GalA-α1,4)-GalA) containing components of the RG-II backbone and its side chains. We showed that the catalytic module of BT1017 adopts an α/β-hydrolase fold, consisting of a central twisted 10-stranded β-sheet sandwiched by several α-helices. This constitutes a new fold for pectin methylesterases, which are predominantly right-handed β-helical proteins. Bioinformatic analyses revealed that the family is dominated by sequences from prominent genera of the human gut microbiota, including Bacteroides and Prevotella. Our re-sults not only highlight the critical role played by this family of enzymes in pectin metabolism but also provide new insights into the molecular basis of the adaptation of B. thetaiotaomicron to the human gut.

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Biochemical evidence that starch breakdown by Bacteroides thetaiotaomicron involves outer membrane starch-binding sites and periplasmic starch-degrading enzymes.

Journal of Bacteriology ◽

10.1128/jb.171.6.3192-3198.1989 ◽

1989 ◽

Vol 171 (6) ◽

pp. 3192-3198 ◽

Cited By ~ 84

Author(s):

K L Anderson ◽

A A Salyers

Keyword(s):

Outer Membrane ◽

Binding Sites ◽

Bacteroides Thetaiotaomicron ◽

Biochemical Evidence ◽

Starch Breakdown ◽

Degrading Enzymes

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The Glycine Lipids ofBacteroides thetaiotaomicronAre Important for Fitness during GrowthIn VivoandIn Vitro

Applied and Environmental Microbiology ◽

10.1128/aem.02157-18 ◽

2018 ◽

Vol 85 (10) ◽

Cited By ~ 2

Author(s):

Alli Lynch ◽

Seshu R. Tammireddy ◽

Mary K. Doherty ◽

Phillip D. Whitfield ◽

David J. Clarke

Keyword(s):

Amino Acid ◽

Gut Microbiome ◽

Molecular Mechanisms ◽

Cellular Membrane ◽

Bacteroides Thetaiotaomicron ◽

Human Gut ◽

Acyltransferase Activity ◽

Content Type ◽

Health And Disease ◽

And Function

ABSTRACTAcylated amino acids function as important components of the cellular membrane in some bacteria. Biosynthesis is initiated by theN-acylation of the amino acid, and this is followed by subsequentO-acylation of the acylated molecule, resulting in the production of the mature diacylated amino acid lipid. In this study, we use both genetics and liquid chromatography-mass spectrometry (LC-MS) to characterize the biosynthesis and function of a diacylated glycine lipid (GL) species produced inBacteroides thetaiotaomicron. We, and others, have previously reported the identification of a gene, namedglsBin this study, that encodes anN-acyltransferase activity responsible for the production of a monoacylated glycine calledN-acyl-3-hydroxy-palmitoyl glycine (or commendamide). In all of theBacteroidalesgenomes sequenced so far, theglsBgene is located immediately downstream from a gene, namedglsA, that is also predicted to encode a protein with acyltransferase activity. We use LC-MS to show that the coexpression ofglsBandglsAresults in the production of GL inEscherichia coli. We constructed a deletion mutant of theglsBgene inB. thetaiotaomicron, and we confirm thatglsBis required for the production of GL inB. thetaiotaomicron. Moreover, we show thatglsBis important for the ability ofB. thetaiotaomicronto adapt to stress and colonize the mammalian gut. Therefore, this report describes the genetic requirements for the biosynthesis of GL, a diacylated amino acid species that contributes to fitness in the human gut bacteriumB. thetaiotaomicron.IMPORTANCEThe gut microbiome has an important role in both health and disease of the host. The mammalian gut microbiome is often dominated by bacteria from theBacteroidales, an order that includesBacteroidesandPrevotella. In this study, we have identified an acylated amino acid, called glycine lipid, produced byBacteroides thetaiotaomicron, a beneficial bacterium originally isolated from the human gut. In addition to identifying the genes required for the production of glycine lipids, we show that glycine lipids have an important role during the adaptation ofB. thetaiotaomicronto a number of environmental stresses, including exposure to either bile or air. We also show that glycine lipids are important for the normal colonization of the murine gut byB. thetaiotaomicron. This work identifies glycine lipids as an important fitness determinant inB. thetaiotaomicronand therefore increases our understanding of the molecular mechanisms underpinning colonization of the mammalian gut by beneficial bacteria.

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Differential Metabolism of Exopolysaccharides from Probiotic Lactobacilli by the Human Gut Symbiont Bacteroides thetaiotaomicron

Applied and Environmental Microbiology ◽

10.1128/aem.00149-15 ◽

2015 ◽

Vol 81 (12) ◽

pp. 3973-3983 ◽

Cited By ~ 30

Author(s):

Alicia Lammerts van Bueren ◽

Aakanksha Saraf ◽

Eric C. Martens ◽

Lubbert Dijkhuizen

Keyword(s):

Gastrointestinal Tract ◽

Carbohydrate Metabolism ◽

Lactobacillus Reuteri ◽

Bacterial Species ◽

Transport Systems ◽

Bacteroides Thetaiotaomicron ◽

Human Gut ◽

Positive Health ◽

Content Type ◽

Commensal Species

ABSTRACTProbiotic microorganisms are ingested as food or supplements and impart positive health benefits to consumers. Previous studies have indicated that probiotics transiently reside in the gastrointestinal tract and, in addition to modulating commensal species diversity, increase the expression of genes for carbohydrate metabolism in resident commensal bacterial species. In this study, it is demonstrated that the human gut commensal speciesBacteroides thetaiotaomicronefficiently metabolizes fructan exopolysaccharide (EPS) synthesized by probioticLactobacillus reuteristrain 121 while only partially degrading reuteran and isomalto/malto-polysaccharide (IMMP) α-glucan EPS polymers.B. thetaiotaomicronmetabolized these EPS molecules via the activation of enzymes and transport systems encoded by dedicated polysaccharide utilization loci specific for β-fructans and α-glucans. Reduced metabolism of reuteran and IMMP α-glucan EPS molecules may be due to reduced substrate binding by components of the starch utilization system (sus). This study reveals that microbial EPS substrates activate genes for carbohydrate metabolism inB. thetaiotaomicronand suggests that microbially derived carbohydrates provide a carbohydrate-rich reservoir forB. thetaiotaomicronnutrient acquisition in the gastrointestinal tract.

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Crystal structure of dipeptidyl peptidase III from the human gut symbiont Bacteroides thetaiotaomicron

PLoS ONE ◽

10.1371/journal.pone.0187295 ◽

2017 ◽

Vol 12 (11) ◽

pp. e0187295 ◽

Cited By ~ 7

Author(s):

Igor Sabljić ◽

Nevenka Meštrović ◽

Bojana Vukelić ◽

Peter Macheroux ◽

Karl Gruber ◽

...

Keyword(s):

Crystal Structure ◽

Dipeptidyl Peptidase ◽

Bacteroides Thetaiotaomicron ◽

Human Gut

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Kin selection explains the evolution of cooperation in the gut microbiota

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2016046118 ◽

2021 ◽

Vol 118 (6) ◽

pp. e2016046118 ◽

Cited By ~ 1

Author(s):

Camille Simonet ◽

Luke McNally

Keyword(s):

Gut Microbiota ◽

Predictive Power ◽

Inclusive Fitness ◽

Evolution Of Cooperation ◽

Community Management ◽

Human Gut ◽

Full Diversity ◽

Gut Microbe ◽

Disease Understanding ◽

Key Parameter

Through the secretion of “public goods” molecules, microbes cooperatively exploit their habitat. This is known as a major driver of the functioning of microbial communities, including in human disease. Understanding why microbial species cooperate is therefore crucial to achieve successful microbial community management, such as microbiome manipulation. A leading explanation is that of Hamilton’s inclusive-fitness framework. A cooperator can indirectly transmit its genes by helping the reproduction of an individual carrying similar genes. Therefore, all else being equal, as relatedness among individuals increases, so should cooperation. However, the predictive power of relatedness, particularly in microbes, is surrounded by controversy. Using phylogenetic comparative analyses across the full diversity of the human gut microbiota and six forms of cooperation, we find that relatedness is predictive of the cooperative gene content evolution in gut-microbe genomes. Hence, relatedness is predictive of cooperation over broad microbial taxonomic levels that encompass variation in other life-history and ecology details. This supports the generality of Hamilton’s central insights and the relevance of relatedness as a key parameter of interest to advance microbial predictive and engineering science.

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Ruminococcus gnavus, a member of the human gut microbiome associated with Crohn’s disease, produces an inflammatory polysaccharide

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1904099116 ◽

2019 ◽

Vol 116 (26) ◽

pp. 12672-12677 ◽

Cited By ~ 54

Author(s):

Matthew T. Henke ◽

Douglas J. Kenny ◽

Chelsi D. Cassilly ◽

Hera Vlamakis ◽

Ramnik J. Xavier ◽

...

Keyword(s):

Crohn’S Disease ◽

Crohn's Disease ◽

Biosynthetic Gene Cluster ◽

Spectroscopic Studies ◽

Major Type ◽

Human Gut ◽

Toll Like Receptor 4 ◽

Gut Microbe ◽

Research Goal ◽

Inflammatory Bowel

A substantial and increasing number of human diseases are associated with changes in the gut microbiota, and discovering the molecules and mechanisms underlying these associations represents a major research goal. Multiple studies associateRuminococcus gnavus, a prevalent gut microbe, with Crohn’s disease, a major type of inflammatory bowel disease. We have found thatR. gnavussynthesizes and secretes a complex glucorhamnan polysaccharide with a rhamnose backbone and glucose sidechains. Chemical and spectroscopic studies indicated that the glucorhamnan was largely a repeating unit of five sugars with a linear backbone formed from three rhamnose units and a short sidechain composed of two glucose units. The rhamnose backbone is made from 1,2- and 1,3-linked rhamnose units, and the sidechain has a terminal glucose linked to a 1,6-glucose. This glucorhamnan potently induces inflammatory cytokine (TNFα) secretion by dendritic cells, and TNFα secretion is dependent on toll-like receptor 4 (TLR4). We also identify a putative biosynthetic gene cluster for this molecule, which has the four biosynthetic genes needed to convert glucose to rhamnose and the five glycosyl transferases needed to build the repeating pentasaccharide unit of the inflammatory glucorhamnan.

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