Peptidoglycan Molecular Requirements Allowing Detection by Nod1 and Nod2

Abstract

Nod1 and Nod2 are mammalian proteins implicated in the intracellular detection of pathogen-associated molecular patterns. Recently, naturally occurring peptidoglycan (PG) fragments were identified as the microbial motifs sensed by Nod1 and Nod2. Whereas Nod2 detects GlcNAc-MurNAc dipeptide (GM-Di), Nod1 senses a unique diaminopimelate-containing GlcNAc-MurNAc tripeptide muropeptide (GM-TriDAP) found mostly in Gram-negative bacterial PGs. Because Nod1 and Nod2 detect similar yet distinct muropeptides, we further analyzed the molecular sensing specificity of Nod1 and Nod2 toward PG fragments. Using a wide array of natural or modified muramyl peptides, we show here that Nod1 and Nod2 have evolved divergent strategies to achieve PG sensing. By defining the PG structural requirements for Nod1 and Nod2 sensing, this study reveals how PG processing and modifications, either by host or bacterial enzymes, may affect innate immune responses. Nod1 and Nod2 are mammalian proteins implicated in the intracellular detection of pathogen-associated molecular patterns. Recently, naturally occurring peptidoglycan (PG) fragments were identified as the microbial motifs sensed by Nod1 and Nod2. Whereas Nod2 detects GlcNAc-MurNAc dipeptide (GM-Di), Nod1 senses a unique diaminopimelate-containing GlcNAc-MurNAc tripeptide muropeptide (GM-TriDAP) found mostly in Gram-negative bacterial PGs. Because Nod1 and Nod2 detect similar yet distinct muropeptides, we further analyzed the molecular sensing specificity of Nod1 and Nod2 toward PG fragments. Using a wide array of natural or modified muramyl peptides, we show here that Nod1 and Nod2 have evolved divergent strategies to achieve PG sensing. By defining the PG structural requirements for Nod1 and Nod2 sensing, this study reveals how PG processing and modifications, either by host or bacterial enzymes, may affect innate immune responses. Peptidoglycan (PG) 1The abbreviations used are: PG, peptidoglycan; MurNAc, N-acetylmuramic acid; GM, GlcNAc-MurNAc; DAP, diaminopimelic acid; MDP, muramyl dipeptide (MurNAc-l-Ala-d-isoGln); M-Di, muramyl dipeptide (MurNAc-l-Ala-d-Glu); DAPamide, meso-DAP (d)-monoamide; TriDAP, DAP tripeptide (l-Ala-γ-d-Glu-meso-DAP); M-TriDAP, meso-DAP-containing muramyl tripeptide; HPLC, high performance liquid chromatography; MALDI-TOF, matrix-assisted laser desorption/ionization time-of-flight; HEK293T, human embryonic kidney 293T cells.1The abbreviations used are: PG, peptidoglycan; MurNAc, N-acetylmuramic acid; GM, GlcNAc-MurNAc; DAP, diaminopimelic acid; MDP, muramyl dipeptide (MurNAc-l-Ala-d-isoGln); M-Di, muramyl dipeptide (MurNAc-l-Ala-d-Glu); DAPamide, meso-DAP (d)-monoamide; TriDAP, DAP tripeptide (l-Ala-γ-d-Glu-meso-DAP); M-TriDAP, meso-DAP-containing muramyl tripeptide; HPLC, high performance liquid chromatography; MALDI-TOF, matrix-assisted laser desorption/ionization time-of-flight; HEK293T, human embryonic kidney 293T cells. is a major constituent of the cell wall of Gram-positive bacteria and consists of glycan chains of alternating N-acetylglucosamine (GlcNAc) and N-acetylmuramic acid (MurNAc) that are cross-linked to each other by short peptides, allowing the formation of a rigid polymer that surrounds the bacterial cell. During bacterial life cycle, peptidoglycan is constantly degraded by specific hydrolases, and newly synthesized subunits are integrated into the polymeric structure, allowing biological processes such as cell division. In Gram-negative bacteria, a thin layer of peptidoglycan is also found in the periplasmic space. Apart from the thickness and the degree of stem peptides cross-linking, an important difference between Gram-positive and Gram-negative peptidoglycans resides in the nature of the third amino acid of the peptides. In Gram-positive bacteria, this amino acid is commonly a lysine, whereas a diaminopimelic acid is found in most Gram-negative bacteria. Extensive analysis from many bacteria has revealed that, in steady state conditions, peptidoglycans from each bacterial strain have somehow fixed proportions of di-, tri-, tetra-, or pentapeptide substituted to the MurNAc sugar moiety. Nod molecules, including Nod1 and Nod2, are a family of intracellular pattern recognition proteins involved in bacterial detection (1Inohara N. Nuñez G. Nat. Rev. Immunol. 2003; 3: 371-380Crossref PubMed Scopus (853) Google Scholar, 2Chamaillard M. Girardin S.E. Viala J. Philpott D.J. Cell. Microbiol. 2003; 5: 581-592Crossref PubMed Scopus (273) Google Scholar). Recently, the bacterial motifs detected by Nod1 and Nod2 have been identified (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar, 4Chamaillard M. Hashimoto M. Horie Y. Masumoto J. Su Q. Saab L. Ogura Y. Kawasaki A. Fukase K. Kusumoto S. Valvano M.A. Foster S.J. Mak T.W. Nuñez G. Inohara N. Nat. Immunol. 2003; 4: 702-707Crossref PubMed Scopus (993) Google Scholar, 5Girardin S.E. Boneca I.G. Viala J. Chamaillard M. Labigne A. Thomas G. Philpott D.J. Sansonetti P.J. J. Biol. Chem. 2003; 278: 8869-8872Abstract Full Text Full Text PDF PubMed Scopus (1860) Google Scholar, 6Inohara N. Ogura Y. Fontalba A. Gutierrez O. Pons F. Crespo J. Fukase K. Inamura S. Kusumoto S. Hashimoto M. Foster S.J. Moran A.P. Fernandez-Luna J.L. Nuñez G. J. Biol. Chem. 2003; 278: 5509-5512Abstract Full Text Full Text PDF PubMed Scopus (1377) Google Scholar). Nod1 and Nod2 both sense peptidoglycan through the detection of different GlcNAc-MurNAc peptide fragments (also known as muropeptides). Nod1 detects the naturally occurring PG degradation product GlcNAc-MurNAc-l-Ala-γ-d-Glu-meso-DAP (GM-TriDAP) (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar). In contrast, Nod2 senses the muramyl dipeptide MurNAc-l-Ala-d-isoGln, also known as MDP (5Girardin S.E. Boneca I.G. Viala J. Chamaillard M. Labigne A. Thomas G. Philpott D.J. Sansonetti P.J. J. Biol. Chem. 2003; 278: 8869-8872Abstract Full Text Full Text PDF PubMed Scopus (1860) Google Scholar, 6Inohara N. Ogura Y. Fontalba A. Gutierrez O. Pons F. Crespo J. Fukase K. Inamura S. Kusumoto S. Hashimoto M. Foster S.J. Moran A.P. Fernandez-Luna J.L. Nuñez G. J. Biol. Chem. 2003; 278: 5509-5512Abstract Full Text Full Text PDF PubMed Scopus (1377) Google Scholar). As a consequence, Nod2 is a general sensor of PGs from Gram-negative and Gram-positive bacteria because MDP corresponds to the minimal motif found in all PGs. In contrast, the PG motif detected by Nod1, GM-TriDAP, is mostly found in PGs from Gram-negative bacteria, suggesting a specific role for Nod1 in innate immune responses against these bacteria. Therefore, specific PG muropeptides could be considered as a new class of pathogen-associated molecular patterns detected intracellularly by Nod molecules. During the course of a bacterial infection, the structure and muropeptide composition of PG is likely to be modified through the action of host and/or bacterial enzymes. For instance, SltY, a bacterial lytic transglycosylase involved in PG processing, is highly up-regulated during the infection of epithelial cells by Shigella flexneri (7Bartoleschi C. Pardini M.C. Scaringi C. Martino M.C. Pazzani C. Bernardini M.C. Cell. Microbiol. 2002; 4: 613-626Crossref PubMed Scopus (35) Google Scholar). On the host side, lytic enzymes such as lysozyme or amidase degrade PG polymers. Therefore, to better understand the role of Nod molecules in innate immune defense following infection by various bacteria, it is necessary to define the PG structural requirements allowing detection by this class of pattern recognition molecules. For this purpose, we have analyzed the sensing specificity of Nod1 and Nod2 toward a large array of PG products and muramyl peptides, either occurring naturally or engineered synthetically. Our results provide new insights into the specific functions of Nod1 and Nod2 in PG detection during bacterial infections. Amino Acid Isomers—The ll and meso isomers of DAP were purified from the Sigma mixture according to van Heijenoort and Bricas (8van Heijenoort J. Bricas E. Bull. Soc. Chim. Fr. 1968; : 2828-2831Google Scholar). Any possible traces of the meso isomer present in the purified ll-DAP preparation were further eliminated by treatment with the DAP decarboxylase (LysA) in a reaction mixture (100 μl) consisting of 100 mm Tris-HCl buffer, pH 7.5, 1 mm EDTA, 1 mm dithiothreitol, 0.1 mm pyridoxal phosphate, 3 mmll-DAP, and pure His6-tagged LysA from Escherichia coli (13 μg). Incubation was performed for 3 h at 37 °C. l-Apm was obtained from the racemic compound according to Berges et al. (9Berges D.A. DeWolf Jr., W.E. Dunn G.L. Grappel S.F. Newman D.J. Taggart J.J. Gilvarg C. J. Med. Chem. 1986; 29: 89-95Crossref PubMed Scopus (47) Google Scholar). The chemical synthesis of the meso-lanthionine and l-allo-cystathionine has been described previously (10Richaud C. Mengin-Lecreulx D. Pochet S. Johnson E.J. Cohen G.N. Marlière P. J. Biol. Chem. 1993; 268: 26827-26835Abstract Full Text PDF PubMed Google Scholar). The purity of the DAP, lanthionine, and cystathionine isomers could be assessed using a high performance liquid chromatography (HPLC) technique after derivatization with the Marfey reagent (11Mengin-Lecreulx D. Michaud C. Richaud C. Blanot D. van Heijenoort J. J. Bacteriol. 1988; 170: 2031-2039Crossref PubMed Google Scholar, 12Mengin-Lecreulx D. Blanot D. van Heijenoort J. J. Bacteriol. 1994; 176: 4321-4327Crossref PubMed Google Scholar). meso-DAP (d)-monoamide (DAPamide) dihydrobromide was prepared by treatment of N,N′-bisbenzyloxycarbonyl-meso-DAP (d)-monoamide (13van Heijenoort J. Bricas E. Nicot C. Bull. Soc. Chim. Fr. 1969; : 2743-2747Google Scholar) with HBr/acetic acid for 1 h. UDP-MurNAc Peptides—The chemical synthesis of UDP-MurNAc has been reported previously (14Blanot D. Auger G. Liger D. van Heijenoort J. Carbohydr. Res. 1994; 252: 107-115Crossref Scopus (20) Google Scholar). UDP-MurNAc-l-Ala was synthesized from UDP-MurNAc using purified E. coli MurC synthetase (15Auger G. Martin L. Bertrand J. Ferrari P. Fanchon E. Vaganay S. Pétillot Y. van Heijenoort J. Blanot D. Dideberg O. Prot. Expr. Purif. 1998; 13: 23-29Crossref PubMed Scopus (46) Google Scholar), UDP-MurNAc-l-Ala-d-Glu was synthesized from UDP-MurNAc-l-Ala using purified E. coli MurD synthetase (16Bertrand J.A. Auger G. Martin L. Fanchon E. Blanot D. Le D. van Heijenoort J. Dideberg O. J. Biol. PubMed Scopus Google Scholar), and and and were as described by et al. Mengin-Lecreulx D. van Heijenoort J. PubMed Scopus Google Scholar). and were synthesized as described previously D. Blanot D. van Heijenoort J. J. Bacteriol. 1994; 176: 4321-4327Crossref PubMed Google Scholar). was synthesized in a reaction mixture (100 μl) consisting of 100 mm Tris-HCl buffer, pH 100 mm mm 1 mm mmll-DAP, and pure His6-tagged from E. coli μg). were used for the synthesis of and with l-Apm and meso-DAP (d)-monoamide Because of different and the from was used to or and were synthesized in a reaction mixture (100 μl) 100 mm Tris-HCl buffer, pH mm mm 1 mm or and pure His6-tagged from S. μg). In all was performed at 37 °C. and were by the specific of the of the by purified of E. according to et al. A. J. PubMed Scopus Google Scholar). In all the were by the of of and the different were purified by a using with mm at a of an at pH for a of pH to was between and The of the was assessed by matrix-assisted laser desorption/ionization found for the and were in and for the and were and MurNAc and and were obtained from by treatment and as described by Michaud et al. C. Blanot D. van Heijenoort J. J. PubMed Scopus Google Scholar). were by acid 100 of the were purified as a mixture of the and by was by treatment of the with at 37 for by with of acid G. van Heijenoort J. Blanot D. J. Chem. Scopus Google Scholar). was purified by a using with acid at a of at was prepared according to and E. J. Chem. Soc. Scopus Google Scholar). The tri-, tetra-, and and is meso-DAP or were obtained by treatment of the with purified E. coli amidase according to the of van Heijenoort et al. Heijenoort J. C. van Heijenoort Y. J. PubMed Scopus Google Scholar). was obtained by treatment of the tripeptide with in a reaction mixture (100 μl) consisting of mm buffer, pH 1 mm and h of at 37 the reaction was as by the of an of and the of a new to the was detected with the amino acid The different peptides were purified by conditions, using Amino Acid acid and amino sugar were with a after of in at for h. were a Foster in the with The were prepared as 1 of a of compound in at was the and with 1 of a of acid in 0.1 and were obtained by from a were in the at an of and an of mixture of and was used as an PG and from E. coli was and purified as described previously (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar). The used for the of polymeric was with lytic enzymes including and purified human amidase as described S. Microbiol. PubMed Scopus Google Scholar). of of purified PG with and amidase were in mm and mm Tris-HCl mm and were at 37 in the of of and of PG muropeptides were by high performance liquid chromatography as described previously (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar). and were in modified to cells were into at a of as described previously S.E. M. Bertin J. P.S. M. Sansonetti P.J. Philpott D.J. PubMed Scopus Google Scholar). and for Nod1 was from Nuñez of and has been described previously N. L. Y. C. C. J. D. J. Nuñez G. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The for Nod2 was from Thomas were in cells as described previously S.E. M. Bertin J. P.S. M. Sansonetti P.J. Philpott D.J. PubMed Scopus Google Scholar). the of by muramyl peptides, and in cells Nod1 or Nod2 were as described by Inohara et al. N. Ogura Y. A. Nuñez G. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). cells were with of Nod1 or of Nod2 of the 1 of PG or of muramyl peptides were to the cell and the was following h of were performed in and at show S.E. of the of Nod1 by from PG was as described previously (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar). PG be degraded by major of lytic enzymes. The class bacterial such as and lysozyme enzymes that the of the between and MurNAc in glycan to the class of PG lytic enzymes that the between the of MurNAc and to Nod1 and Nod2 could detect PG following by or this PG from E. coli was The of polymeric PG to or epithelial cells the and of the as described the E. coli PG preparation and that the could the Nod1 or Nod2 that the PG motifs Nod1 and Nod2 are present in the polymeric PG The was to PG previously by amidase or In this the in the of and peptides after amidase that of PG muropeptides that are detected both by Nod1 and Nod2 a that was because Nod1 and Nod2 detect and amidase of fragments that were detected by Nod1 by Nod2 PG from E. coli with was using at In such conditions, the MurNAc are and structure, whereas the amino that the chemical of the PG sugar Nod2 sensing, whereas Nod1 could detect both and PG fragments these results that Nod1 and Nod2 different toward PG sensing. Nod1 sensing to the PG Nod2 an MurNAc substituted to a these results that either by the host or the bacteria, could affect the between and PG sensing. define the minimal PG motif detected by Nod1 and Nod2, muramyl peptides, peptides, peptides, and peptides were or and purity was by including and amino acid analysis also and the muramyl dipeptide used in this study is in as whereas the muramyl dipeptide that was used in by and is in the amino acid is an to MDP, because is in PGs and because the other muramyl peptides of this study also molecules were analyzed for to the and that Nod1 detects detects results (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar) that that Nod1 detects a naturally occurring muropeptide with a GM-TriDAP, and further show that the sugar found in a role in Nod1 the tripeptide Nod1 as as PG is detected by Nod1 peptides and and amino and and that the minimal motif detected by Nod1 is dipeptide is sensed by Nod1 with in suggesting that the present in may be for detection of PG by of and meso-DAP is in Nod1, that the between these amino is for Nod1 sensing. these results that Nod1 detects PGs the motif in the meso-DAP is in are in with a study from M. Hashimoto M. Horie Y. Masumoto J. Su Q. Saab L. Ogura Y. Kawasaki A. Fukase K. Kusumoto S. Valvano M.A. Foster S.J. Mak T.W. Nuñez G. Inohara N. Nat. Immunol. 2003; 4: 702-707Crossref PubMed Scopus (993) Google Scholar) that used a of chemical synthesis to that is the minimal PG motif detected by and meso-DAP amino are from and a of bacterial we also that Nod1 detects this compound the motif with a meso-DAP analysis revealed that is sensed by Nod1 with GM-TriDAP, M-TriDAP, or in and the for this of these could be detected by Nod2. As of the molecules the MurNAc sugar could Nod2 In with results (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar), we that Nod2 detects muramyl dipeptide in this MDP in 5Girardin S.E. Boneca I.G. Viala J. Chamaillard M. Labigne A. Thomas G. Philpott D.J. Sansonetti P.J. J. Biol. Chem. 2003; 278: 8869-8872Abstract Full Text Full Text PDF PubMed Scopus (1860) Google Scholar) detect or the that Nod2 detects as as MDP that of the amino acid the of Nod2 to detect muramyl The with a role for a meso-DAP in detection of of the meso-DAP that are for Nod1 sensing, we purified muramyl in the nature of the third amino acid and for to In to M-TriDAP, the muramyl tripeptide detected by Nod1 is the is naturally present in PGs from Gram-negative bacteria such as meso-lanthionine from meso-DAP by the of the by a the other muramyl were sensed by Nod1 the of the the substituted the because and muramyl peptide naturally present in PGs from bacteria such as are detected by the of the because the of meso-lanthionine by an amino acid that could be in E. coli PG (10Richaud C. Mengin-Lecreulx D. Pochet S. Johnson E.J. Cohen G.N. Marlière P. J. Biol. Chem. 1993; 268: 26827-26835Abstract Full Text PDF PubMed Google Scholar, 12Mengin-Lecreulx D. Blanot D. van Heijenoort J. J. Bacteriol. 1994; 176: 4321-4327Crossref PubMed Google Scholar), to a muramyl tripeptide that is detected with by the amino substituted the because of meso-DAP by detection by Nod1 of the muramyl tripeptide and the of the the because is detected by with the results also these results the of each of meso-DAP for PG detection by Nod1, the amino and the amino and amino be into a peptide with acid and the be because Nod1 detect and the of the muramyl were also for detection by Nod2. we that Nod2 detects also and results the following specific of sensing of PG that from of Nod2 detects muramyl peptides with a peptide of dipeptide or tripeptide the third amino acid is to a The Nod2 sensing, because Nod2 detects the of the by a the of Nod2 sensing, suggesting that the of sensing by Nod2 chemical of the The of the third amino is for detection of PG, because Nod2 detects and with In the of the detection of PG by Nod1, an could be identified between (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar) and results by M. Hashimoto M. Horie Y. Masumoto J. Su Q. Saab L. Ogura Y. Kawasaki A. Fukase K. Kusumoto S. Valvano M.A. Foster S.J. Mak T.W. Nuñez G. Inohara N. Nat. Immunol. 2003; 4: 702-707Crossref PubMed Scopus (993) Google Scholar). in we of Nod1 by PG, of S. flexneri PG were (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar). is with the present that Nod1 detect a muramyl peptide found in PG In contrast, Inohara and reported of Nod1 by PG M. Hashimoto M. Horie Y. Masumoto J. Su Q. Saab L. Ogura Y. Kawasaki A. Fukase K. Kusumoto S. Valvano M.A. Foster S.J. Mak T.W. Nuñez G. Inohara N. Nat. Immunol. 2003; 4: 702-707Crossref PubMed Scopus (993) Google Scholar). this we analyzed the detection of PG by Nod1 in further that, whereas 1 PG could Nod1 as we reported previously (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar), the PG to a of Nod1 In contrast, S. PG is to Nod1 By with PG from a Gram-negative such as S. we that PG is to Nod1 to a similar and muropeptides from PG by was for of Nod1 corresponds to was a to the could Nod1 Therefore, a high of PG is of muropeptides DAP, of DAP are also found in has been reported previously by Foster and and the have that PG and A. G. G. Foster S.J. J. Bacteriol. PubMed Google Scholar). these results that PG is a of Nod1 as with Gram-negative bacterial because the DAP acid is to a high During bacterial PG are synthesized in a highly in specific enzymes the of amino to the muramyl During this the MurNAc sugar is through to an to the of a to the could affect detection of muramyl peptides by Nod1 or Nod2. this UDP-MurNAc peptides of different are in Gram-negative or Gram-positive bacterial were or as because the amino of the pentapeptide are as a to a by the Nod1 detected both and that the of a to the MurNAc affect the of Nod1 to detect was because the sugar is for Nod1 to achieve PG sensing. In the affect the of Nod2 to detect or or Nod2 detect M-TriDAP, it senses as as MDP or how the of an to the MurNAc detection by Nod2, from and were modified to the peptides and Using these we could that the to the of MurNAc is to Nod2 to muramyl peptide we that these may with the present the meso-DAP the of a the of Nod2 to sense and these results show that the of a the MurNAc sugar sensing of muramyl peptides. has been in the of the role of intracellular bacterial sensing by Nod molecules in innate Nod1 a role as a epithelial the of defense against because epithelial cells from are to bacterial products into (3Girardin S.E. Boneca I.G. Carneiro L.A. Antignac A. Jéhanno M. Viala J. Tedin K. Taha M.-K. Labigne A. Zaehringer U. Coyl A.J. Di Stefano P.S. Bertin J. Sansonetti P.J. Philpott D.J. Science. 2003; 300: 1584-1587Crossref PubMed Scopus (1225) Google Scholar). In the of Nod2, has that with in Nod2 a to a by a of the Chamaillard M. S. J. S. C. M. Y. A. P. C. J. M. Thomas G. PubMed Scopus Google Scholar, Y. Inohara N. Moran Nuñez G. PubMed Scopus Google Scholar). in Nod2 could be with in PG or muramyl dipeptide sensing (5Girardin S.E. Boneca I.G. Viala J. Chamaillard M. Labigne A. Thomas G. Philpott D.J. Sansonetti P.J. J. Biol. Chem. 2003; 278: 8869-8872Abstract Full Text Full Text PDF PubMed Scopus (1860) Google Scholar, 6Inohara N. Ogura Y. Fontalba A. Gutierrez O. Pons F. Crespo J. Fukase K. Inamura S. Kusumoto S. Hashimoto M. Foster S.J. Moran A.P. Fernandez-Luna J.L. Nuñez G. J. Biol. Chem. 2003; 278: 5509-5512Abstract Full Text Full Text PDF PubMed Scopus (1377) Google Scholar, M. Philpott D. Girardin S.E. S. F. M. Zaehringer U. Sansonetti P.J. Thomas G. U. S. A. 2003; PubMed Scopus Google Scholar). The of the PG structural requirements allowing detection is a toward the specific of Nod1 and Nod2 in an innate immune defense against bacteria. we have that Nod1 and Nod2 detect both PG degradation products and Our results have identified as the of a compound detected by both Nod1 and Nod2, suggesting the of an of Nod1 and Nod2 in the detection of PG products from Gram-negative bacteria. results that either by bacteria or the the toward sensing, may the host to Recently, the nature of the mammalian amidase has been identified as E. C. J. Res. 2003; PubMed Scopus Google Scholar). that a role in the specific of Nod1 at the of Nod2. we have that Nod1 is to detect PGs from Gram-negative bacteria of Gram-negative bacteria such as and a of meso-DAP in The of may an important for the of these because bacteria the composition of PG during the muropeptide detected by Nod1 and Nod2 for the of PG be with immune for purified E. coli