Publications

• Phosphorylation of muramyl peptides by NAGK is required for NOD2 activation.

  Stafford C.A., Gassauer A.M., de Oliveira Mann C.C., Tanzer M.C., Fessler E., Wefers B., Nagl D., Kuut G., Sulek K., Vasilopoulou C., Schwojer S.J., Wiest A., Pfautsch M.K., Wurst W., Yabal M., Froehlich T., Mann M., Gisch N., Jae L.T., Hornung V.

  Bacterial cell wall components provide various unique molecular structures that are detected by pattern recognition receptors (PRRs) of the innate immune system as non-self. Most bacterial species form a cell wall that consists of peptidoglycan (PGN), a polymeric structure comprising alternating a ... >> More

  Bacterial cell wall components provide various unique molecular structures that are detected by pattern recognition receptors (PRRs) of the innate immune system as non-self. Most bacterial species form a cell wall that consists of peptidoglycan (PGN), a polymeric structure comprising alternating amino sugars that form strands cross-linked by short peptides. Muramyl dipeptide (MDP) has been well documented as a minimal immunogenic component of peptidoglycan^1-3. MDP is sensed by the cytosolic nucleotide-binding oligomerization domain-containing protein 2⁴ (NOD2). Upon engagement, it triggers pro-inflammatory gene expression, and this functionality is of critical importance in maintaining a healthy intestinal barrier function⁵. Here, using a forward genetic screen to identify factors required for MDP detection, we identified N-acetylglucosamine kinase (NAGK) as being essential for the immunostimulatory activity of MDP. NAGK is broadly expressed in immune cells and has previously been described to contribute to the hexosamine biosynthetic salvage pathway⁶. Mechanistically, NAGK functions upstream of NOD2 by directly phosphorylating the N-acetylmuramic acid moiety of MDP at the hydroxyl group of its C6 position, yielding 6-O-phospho-MDP. NAGK-phosphorylated MDP-but not unmodified MDP-constitutes an agonist for NOD2. Macrophages from mice deficient in NAGK are completely deficient in MDP sensing. These results reveal a link between amino sugar metabolism and innate immunity to bacterial cell walls. << Less

  Nature 609:590-596(2022) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• Structural and biochemical insights into the molecular mechanism of N-acetylglucosamine/N-Acetylmuramic acid kinase MurK from Clostridium acetobutylicum.

  He X., Liu C., Li X., Yang Q., Niu F., An L., Fan Y., Li Y., Zhou Z., Zhou H., Yang X., Liu X.

  MurK is a MurNAc- and GlcNAc-specific amino sugar kinase, phosphorylates MurNAc and GlcNAc at the 6-hydroxyl group in an ATP-dependent manner, and contributes to the recovery of both amino sugars during the cell wall turnover in Clostridium acetobutylicum. Herein, we determined the crystal structu ... >> More

  MurK is a MurNAc- and GlcNAc-specific amino sugar kinase, phosphorylates MurNAc and GlcNAc at the 6-hydroxyl group in an ATP-dependent manner, and contributes to the recovery of both amino sugars during the cell wall turnover in Clostridium acetobutylicum. Herein, we determined the crystal structures of MurK in complex with MurNAc, GlcNAc, and glucose, respectively. MurK represents the V-shaped fold, which is divided into a small N-terminal domain and a large C-terminal domain. The catalytic pocket is located within the deep cavity between the two domains of the MurK monomer. We mapped the significant enzyme-substrate interactions, identified key residues involved in the catalytic activity of MurK, and found that residues Asp⁷⁷ and Arg⁷⁸ from the β4-α2-loop confer structural flexibilities to specifically accommodate GlcNAc and MurNAc, respectively. Moreover, structural comparison revealed that MurK adopts closed-active conformation induced by the N-acetyl moiety from GlcNAc/MurNAc, rather than closed-inactive conformation induced by glucose, to carry out its catalytic reaction. Taken together, our study provides structural and functional insights into the molecular mechanism of MurK for the phosphorylation of both MurNAc and GlcNAc, sugar substrate specificity, and conformational changes upon sugar substrate binding. << Less

  Int J Biol Macromol 280:135747-135747(2024) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Characterization of an N-acetylmuramic acid/N-acetylglucosamine kinase of Clostridium acetobutylicum.

  Reith J., Berking A., Mayer C.

  We report here the cloning and characterization of a cytoplasmic kinase of Clostridium acetobutylicum, named MurK (for murein sugar kinase). The enzyme has a unique specificity for both amino sugars of the bacterial cell wall, N-acetylmuramic acid (MurNAc) and N-acetylglucosamine (GlcNAc), which a ... >> More

  We report here the cloning and characterization of a cytoplasmic kinase of Clostridium acetobutylicum, named MurK (for murein sugar kinase). The enzyme has a unique specificity for both amino sugars of the bacterial cell wall, N-acetylmuramic acid (MurNAc) and N-acetylglucosamine (GlcNAc), which are phosphorylated at the 6-hydroxyl group. Kinetic analyses revealed Km values of 190 and 127 μM for MurNAc and GlcNAc, respectively, and a kcat value (65.0 s(-1)) that was 1.5-fold higher for the latter substrate. Neither the non-N-acetylated forms of the cell wall sugars, i.e., glucosamine and/or muramic acid, nor epimeric hexoses or 1,6-anhydro-MurNAc were substrates for the enzyme. MurK displays low overall amino acid sequence identity (24%) with human GlcNAc kinase and is the first characterized bacterial representative of the BcrAD/BadFG-like ATPase family. We propose a role of MurK in the recovery of muropeptides during cell wall rescue in C. acetobutylicum. The kinase was applied for high-sensitive detection of the amino sugars in cell wall preparations by radioactive phosphorylation. << Less

  J. Bacteriol. 193:5386-5392(2011) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.