Sialidases and fucosidases of Akkermansia muciniphila are crucial for growth on mucin and nutrient sharing with mucus-associated gut bacteria

Abstract

The mucolytic human gut microbiota specialist Akkermansia muciniphila is proposed to boost mucin-secretion by the host, thereby being a key player in mucus turnover. Mucin glycan utilization requires the removal of protective caps, notably fucose and sialic acid, but the enzymatic details of this process remain largely unknown. Here, we describe the specificities of ten A. muciniphila glycoside hydrolases, which collectively remove all known sialyl and fucosyl mucin caps including those on double-sulfated epitopes. Structural analyses revealed an unprecedented fucosidase modular arrangement and explained the sialyl T-antigen specificity of a sialidase of a previously unknown family. Cell-attached sialidases and fucosidases displayed mucin-binding and their inhibition abolished growth of A. muciniphila on mucin. Remarkably, neither the sialic acid nor fucose contributed to A. muciniphila growth, but instead promoted butyrate production by co-cultured Clostridia. This study brings unprecedented mechanistic insight into the initiation of mucin O-glycan degradation by A. muciniphila and nutrient sharing between mucus-associated bacteria.

Mechanism of 2’-Fucosyllactose degradation by Human-AssociatedAkkermansia

“…In our phylogenetic analysis, genes from clade 1 were significantly upregulated in both species. Interestingly, this protein in A. muciniphila Muc T , Amuc_0846, has weak activity on 2'-FL but strong mucin binding activity (32). Also in agreement, both strains downregulated (significantly only for A. biwaensis another GH29 belonging to clade 7.…”

“…Despite this genomic difference, both A. muciniphila Muc T and A. biwaensis CSUN-19 grew in 2'-FL in media lacking mucin. Furthermore, both strains significantly induced the expression of homologs of a gene encoding a GH29 fucosidase (clade 1), for which the purified protein from A. muciniphila has previously been shown to have weak activity against para-nitrophenyl-α-L-fucoside, an analog of 2'-FL (32). Interestingly, a gene unique to A. biwaensis strains encoding a different GH29 was also significantly upregulated on 2'-FL.…”

“…Analysis of putative fucosidase and 𝛽-galactosidase genes across Akkermansia species revealed differences in the number and diversity of GH29, GH95, and GH2 genes that was phylogroup specific. Recently, six fucosidases from A. muciniphila Muc T have been characterized, demonstrating each enzyme has specific activity on different fucosylated Growth of Akkermansia on 2'-FL 22 substrates that are diet-or host-derived (32). These corroborate previous work demonstrating that members of the GH95 family hydrolyze 1,2-ɑ-L-fucosidase linkages, while the other dominant fucosidase family, GH29, is divided into two subfamilies in which Group A has little substrate specificity and GH29 Group B hydrolyzes ɑ-1,3/1,4-L-fucosidase linkages (63,64).…”

“…Although Akkermansia are predominantly mucin-degrading specialists, given the structural and compositional similarities between HMOs and mucin oligosaccharides, species may use similar enzyme to break down HMOs (26,32). Most of the efforts in developing a mechanistic understanding of Akkermansia utilization of host glycans thus far have centered around a single isolate of A. muciniphila (Muc T ).…”

“…Most of the efforts in developing a mechanistic understanding of Akkermansia utilization of host glycans thus far have centered around a single isolate of A. muciniphila (Muc T ). The Muc T strain has been shown to use a diversity of GHs including fucosidases, 𝛽-hexosaminidases, and 𝛽-galactosidases for degrading a variety of human-glycans including HMOs and lactose (26,32,33). Recently, ourselves and others have identified and isolated at least four species-level phylogroups (AmI-AmIV) of human associated Akkermansia.…”