Elucidation of Akkermansia muciniphila Probiotic Traits Driven by Mucin Depletion

Shin, Jisoo; Noh, Jung‐Ran; Chang, Dong-Ho; Kim, Yong‐Hoon; Kim, Myung Hee; Lee, Eaum Seok; Cho, Suhyung; Ku, Bon Jeong; Rhee, Moon-Soo; Kim, Byoung-Chan; Lee, Chul‐Ho; Cho, Byung-Kwan

doi:10.3389/fmicb.2019.01137

Cited by 88 publications

(59 citation statements)

References 53 publications

(99 reference statements)

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“…Akkermansia muciniphila can use mucus as the sole source of carbon and nitrogen by producing several mucolytic enzymes [33,41]. Therefore, Akkermansia muciniphila also plays an important role in mucin degradation [42]. However, the mechanisms how Akkermansia muciniphila degrades mucins are still poorly understood.…”

Section: Introductionmentioning

confidence: 99%

A Purified Aspartic Protease from Akkermansia Muciniphila Plays an Important Role in Degrading Muc2

Meng

Wang

Lan

et al. 2019

IJMS

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Akkermansia muciniphila can produce various mucin-degrading proteins. However, the functional characteristics of these proteins and their role in mucin degradation are unclear. Of the predicted protein-coding genes, Amuc_1434, which encodes for a hypothetical protein, is the focus in this study. A recombinant enzyme Amuc_1434 containing the 6× His-tag produced in Escherichia coli (hereinafter termed Amuc_1434*) was isolated to homogeneity and biochemically characterised. Results showed that the enzyme can hydrolyse hemoglobin with an activity of 17.21 U/μg. The optimal pH and temperature for hemoglobin hydrolysis of Amuc_1434* were found to be around 8.0 and 40 °C, respectively. Amuc_1434* is identified as a member of the aspartic protease family through the action of inhibitor pepstatin A. Amuc_1434* promotes the adhesion of colon cancer cell line LS174T, which can highly express Muc2. Significantly Amuc_1434* can degrade Muc2 of colon cancer cells. Amuc_1434 is mainly located in the colon of BALB/c mice. These results suggest that the presence of Amuc_1434 from Akkermansia muciniphila may be correlated with the restoration of gut barrier function by decreasing mucus layer thickness.

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Section: Introductionmentioning

confidence: 99%

A Purified Aspartic Protease from Akkermansia Muciniphila Plays an Important Role in Degrading Muc2

Meng

Wang

Lan

et al. 2019

IJMS

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“…Inspection of previously published transcriptomic and proteomic data from four Gram-negative prominent mucin degrading HGM species ( B. thetaiotaomicron, B. fragilis, B. caccae and A. mucinphila ) identified genes and proteins which are likely involved in mucin breakdown 8,11,19–22 . These included many exo-acting enzymes from CAZy families (carbohydrate active enzymes; CAZymes) previously identified as involved degradation of O-glycans, and in the case of Bacteroides spp., SusCD glycan import apparatus and putative surface glycan binding proteins (pSGBPs; Supplementary Fig.…”

Section: Resultsmentioning

confidence: 99%

Prominent members of the human gut microbiota express endo-acting O-glycanases to initiate mucin breakdown

Crouch

Liberato

Urbanowicz

et al. 2019

Preprint

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33The human gut microbiota (HGM) are closely associated with health, development and 34 disease. The thick intestinal mucus layer, especially in the colon, is the key barrier between 35 the contents of the lumen and the epithelial cells, providing protection against infiltration by 36 the microbiota as well potential pathogens. The upper layer of the colonic mucus is a niche for 37 a subset of the microbiota which utilise the mucin glycoproteins as a nutrient source and mucin 38 grazing by the microbiota appears to play a key role in maintaining barrier function as well as 39 community stability. Despite the importance of mucin breakdown for gut health, the 40 mechanisms by which gut bacteria access this complex glycoprotein are not well understood. 41The current model for mucin degradation involves exclusively exo-acting glycosidases that 42 sequentially trim monosaccharides from the termini of the glycan chains to eventually allow 43 access to the mucin peptide backbone by proteases. However, this model is in direct contrast 44 to the Sus paradigm of glycan breakdown used by the Bacteroidetes which involves 45 extracellular cleavage of glycans by surface located endo-acting enzymes prior to import of 46 the oligosaccharide products. Here we describe the discovery and characterisation of endo-47 acting family 16 glycoside hydrolases (GH16s) from prominent mucin degrading gut bacteria 48 that specifically target the oligosaccharide side chains of intestinal mucins from both animals 49 and humans. These endo-acting O-glycanases display β1,4-glactosidase activity and in 50 several cases are surface located indicating they are involved in the initial step in mucin 51 breakdown. The data suggest a new paradigm for mucin breakdown by the microbiota and 52 the endo-mucinases provide a potential tool to explore changes that occur in mucin structure 53 in intestinal disorders such as inflammatory bowel disease and colon cancer. 54 suggesting endo-like cleavage of the O-glycan chains. To investigate the identity of these 126 products in more detail, the products were labelled with the fluorophore procainamide and 127 analysed by liquid chromatography-fluorescence-detection-electrospray-mass spectrometry 128 (LC-FLD-ESI-MS) and the glycan structures determined by MS/MS (Fig. 2a). The data show 129 that oligosaccharides are produced by the GH16 enzymes with between 2 and 6 alternating 130 hexose and HexNAc sugars that are likely to be sections of the polyLacNAc repeats that 131 form the repeating unit of O-glycan chains (Fig. 1b). The reducing ends were all hexoses, 132 11 Desai, M. S. et al. A Dietary Fiber-Deprived Gut Microbiota Degrades the Colonic Mucus 595 Barrier and Enhances Pathogen Susceptibility. Egan, M. et al. Cross-feeding by Bifidobacterium breve UCC2003 during co-cultivation with 598Bifidobacterium bifidum PRL2010 in a mucin-based medium.

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“…Akkermansia muciniphila is a key component of the gut microbiome with many biological functions, which can regulate the level of Muc2 through its own proteolytic enzyme [53][54][55]. Previous studies done by our group demonstrated that that Amuc_1434* has the ability to degrade Muc2 and is mainly expressed in the colon in mice.…”

Section: Discussionmentioning

confidence: 99%

Akkermansia muciniphila Aspartic Protease Amuc_1434* Inhibits Human Colorectal Cancer LS174T Cell Viability via TRAIL-Mediated Apoptosis Pathway

Meng

Zhang

et al. 2020

IJMS

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Mucin2 (Muc2) is the main component of the intestinal mucosal layer and is highly expressed in mucous colorectal cancer. Previous studies conducted by our lab found that the recombinant protein Amuc_1434 (expressed in Escherichia coli prokaryote cell system, hereinafter termed Amuc_1434*), derived from Akkermansia muciniphila, can degrade Muc2. Thus, the main objective of this study was to explore the effects of Amuc_1434* on LS174T in colorectal cancer cells expressing Muc2. Results from this study demonstrated that Amuc_1434* inhibited the proliferation of LS174T cells, which was related to its ability to degrade Muc2. Amuc_1434* also blocked the G0/G1 phase of the cell cycle of LS174T cells and upregulated the expression of tumor protein 53 (p53), which is a cell cycle-related protein. In addition, Amuc_1434* promoted apoptosis of LS174T cells and increased mitochondrial ROS levels in LS174T cells. The mitochondrial membrane potential of LS174T cells was also downregulated by Amuc_1434*. Amuc_1434* can activate the death receptor pathway and mitochondrial pathway of apoptosis by upregulating tumor-necrosis-factor-related apoptosis-inducing ligand (TRAIL). In conclusion, our study was the first to demonstrate that the protein Amuc_1434* derived from Akkermansia muciniphila suppresses LS174T cell viability via TRAIL-mediated apoptosis pathway.

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Elucidation of Akkermansia muciniphila Probiotic Traits Driven by Mucin Depletion

Cited by 88 publications

References 53 publications

A Purified Aspartic Protease from Akkermansia Muciniphila Plays an Important Role in Degrading Muc2

A Purified Aspartic Protease from Akkermansia Muciniphila Plays an Important Role in Degrading Muc2

Prominent members of the human gut microbiota express endo-acting O-glycanases to initiate mucin breakdown

Akkermansia muciniphila Aspartic Protease Amuc_1434* Inhibits Human Colorectal Cancer LS174T Cell Viability via TRAIL-Mediated Apoptosis Pathway

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