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. 2024 May 21;90(5):e0004624.
doi: 10.1128/aem.00046-24. Epub 2024 Apr 2.

Alginate oligosaccharide assimilation by gut microorganisms and the potential role in gut inflammation alleviation

Xiao-Qing Ma #  1 , Bing Wang #  1 , Wei Wei #  2 , Fang-Cheng Tan  1 , Hang Su  1 , Jun-Zhe Zhang  1 , Chen-Yang Zhao  2 , Hua-Jun Zheng  3 , Yan-Qin Feng  4 , Wei Shen  4 , Jin-Bo Yang  2 , Fu-Li Li  1   5   6
Affiliations

Alginate oligosaccharide assimilation by gut microorganisms and the potential role in gut inflammation alleviation

Xiao-Qing Ma et al. Appl Environ Microbiol. .

Abstract

Dietary fiber metabolism by gut microorganisms plays important roles in host physiology and health. Alginate, the major dietary fiber of daily diet seaweeds, is drawing more attention because of multiple biological activities. To advance the understanding of alginate assimilation mechanism in the gut, we show the presence of unsaturated alginate oligosaccharides (uAOS)-specific alginate utilization loci (AUL) in human gut microbiome. As a representative example, a working model of the AUL from the gut microorganism Bacteroides clarus was reconstructed from biochemistry and transcriptome data. The fermentation of resulting monosaccharides through Entner-Doudoroff pathway tunes the metabolism of short-chain fatty acids and amino acids. Furthermore, we show that uAOS feeding protects the mice against dextran sulfate sodium-induced acute colitis probably by remodeling gut microbiota and metabolome.

Importance: Alginate has been included in traditional Chinese medicine and daily diet for centuries. Recently discovered biological activities suggested that alginate-derived alginate oligosaccharides (AOS) might be an active ingredient in traditional Chinese medicine, but how these AOS are metabolized in the gut and how it affects health need more information. The study on the working mechanism of alginate utilization loci (AUL) by the gut microorganism uncovers the role of unsaturated alginate oligosaccharides (uAOS) assimilation in tuning short-chain fatty acids and amino acids metabolism and demonstrates that uAOS metabolism by gut microorganisms results in a variation of cell metabolites, which potentially contributes to the physiology and health of gut.

Keywords: Bacteroides clarus; alginate lyase; alginate utilization loci; dietary fiber; gut microbiome.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig 1
Fig 1
The human gut microbiome harbors alginate utilization loci. (A) Distribution of predicted alginate lyases from the HMP database (see Data Set S1A for more details). (B) Genomic organization of AUL in different species. The representative genome ID of B. clarus YIT 12056 is AFBM00000000.1. Genes encoding proteins of known or predicted functions are color-classified.
Fig 2
Fig 2
Unsaturated alginate oligosaccharides support growth of B. clarus. (A) The highest OD600nm of B. clarus grown on glucose, unsaturated AOS, saturated AOS, polyM, polyG, and polyMG blocks for 48 h. (B) Growth curves of B. clarus on uAOS and glucose. (D) TLC analysis of residual oligosaccharides in culture medium.
Fig 3
Fig 3
BcGntR regulated the bidirectional transcription of the AUL. (A) Schematic diagram of the two promoter areas. (B) EMSAs of BcGntR binding to promoter1 and promoter2. The shift of promoter DNA was affected by increasing concentrations of BcGntR. The addition of non-labeled promoter DNA led to competitive binding with the labeled fragment, indicating specific binding. (C) BcGntR inhibited the activity of promoter1 and promoter2 in E. coli. The constitutive expression of GFP controlled by promoter1 and promoter2 was detected by western blot in E. coli. The simultaneous expression of BcGntR inhibited the expression of GFP. The GFP encoding sequence following promoter1 or promoter2 was cloned into puc57 vector. The BcGntR expression cassette was cloned downstream of the gfp gene. T7 operator was used to mediate the expression of BcGntR. (D) The binding activity of BcGntR to the promoter of AUL in the presence of different saccharides. The unsaturated alginate disaccharide eliminated the binding of BcGntR with the promoter.
Fig 4
Fig 4
Depolymerization mechanism of uAOS by alginate lyase BcAlyPL6 and BcAlyPL17. (A) Degradation of polyuronate blocks by two exolytic alginate lyases. The released products were monitored by ultraviolet detection at 235 nm. Unsaturated monosaccharides were spontaneously converted into 4-deoxy-l-erythro-5-hexoseulose uronic acid which led to the decrease of A235. (B) Cooperative degradation of PolyMG blocks by BcAlyPL6 and BcAlyPL17. (C) Western blot detection of protein location. rAlyPL6 or rAlyPL17, recombinant protein expressed in E. coli. WCE, whole-cell extraction; PK-treated, proteinase K-treated cells; OM, outer membrane; IM, inner membrane. (D) Extracellular and intracellular alginate lyase activity of B. clarus grown on uAOS. AOS was used as the substrate and degradation samples were taken at indicated time. TLC was used to show the degradation products. M, marker; extra, degradation by extracellular fraction; intra, degradation by intracellular enzymes; mono, monosaccharide; di, disaccharides; DEH, 4-deoxy-l-ery thro-5-hexoseulose uronic acid.
Fig 5
Fig 5
Deacetylating activity of BcAae. (A) Acetylation degree of acetylalginate substrate was measured after incubation with BcAae. (B) Relative activity of the endolytic alginate lyase AlgAT0 toward untreated/BcAae-treated acetylalginate.
Fig 6
Fig 6
A working model of the AUL in B. clarus.
Fig 7
Fig 7
Hypothetical pathway involving genes regulated by uAOS utilization based on the RNA-seq data. ED pathway is involved in the fermentation of uronates. Pyruvate is either turned into acetyl-CoA which is further converted into acetate or turned into lactate by 2-hydroxyacid dehydrogenase consuming NADH. Several amino acid metabolic pathways were also highly upregulated. Locus tag numbers HMPREF9445_RSXXXXX are abbreviated with the last numbers after the “RS.”
Fig 8
Fig 8
uAOS protected mice from DSS-induced colitis through remodeling gut microbiota and metabolome. (A) Schematic diagram of the experimental procedure. The weight loss (B) and clinical score (C) were monitored. (D) Linear discriminate analysis effect size was performed to determine the biomarkers at the genus and species level. The threshold of LDA score was 2.0. (E) Heatmap of changed stool metabolites under different treatments. The statistical analyses were performed using two-way ANOVA in (B) and (C) and one-way ANOVA in (E). The data in (B) and (C) were presented as means ± SEM and P < 0.05 is indicated by # for the comparison between uAOS + DSS and DSS groups, and by * for the comparison between DSS and the control group. In (E), the metabolites restored by uAOS are indicated by * and those significantly induced by uAOS are indicated by #.
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