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An intermolecular binding mechanism involving multiple LysM domains mediates carbohydrate recognition by an endopeptidase.

Wong JE, Midtgaard SR, Gysel K, Thygesen MB, Sørensen KK, Jensen KJ, Stougaard J, Thirup S, Blaise M - Acta Crystallogr. D Biol. Crystallogr. (2015)

Bottom Line: The crystal structure and small-angle X-ray scattering solution studies of this endopeptidase revealed the presence of a homodimer.The structure of the two LysM domains co-crystallized with N-acetyl-chitohexaose revealed a new intermolecular binding mode that may explain the differential interaction between LysM domains and short or long chitin oligomers.By combining the structural information with the three-dimensional model of peptidoglycan, a model suggesting how protein dimerization enhances the recognition of peptidoglycan is proposed.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Carbohydrate Recognition and Signalling, Department of Molecular Biology and Genetics, Aarhus University, Gustav Wieds Vej 10C, 8000 Aarhus, Denmark.

ABSTRACT
LysM domains, which are frequently present as repetitive entities in both bacterial and plant proteins, are known to interact with carbohydrates containing N-acetylglucosamine (GlcNAc) moieties, such as chitin and peptidoglycan. In bacteria, the functional significance of the involvement of multiple LysM domains in substrate binding has so far lacked support from high-resolution structures of ligand-bound complexes. Here, a structural study of the Thermus thermophilus NlpC/P60 endopeptidase containing two LysM domains is presented. The crystal structure and small-angle X-ray scattering solution studies of this endopeptidase revealed the presence of a homodimer. The structure of the two LysM domains co-crystallized with N-acetyl-chitohexaose revealed a new intermolecular binding mode that may explain the differential interaction between LysM domains and short or long chitin oligomers. By combining the structural information with the three-dimensional model of peptidoglycan, a model suggesting how protein dimerization enhances the recognition of peptidoglycan is proposed.

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Structure comparison of the catalytic domain of P60_tth with YkfC from B. cereus. (a) Primary-sequence alignment of P60_tth with YkfC from B. cereus. The secondary structures of the two proteins are indicated. The red sphere indicates the catalytic triad, while the blue spheres indicate other residues that are predicted to be involved in forming the catalytic site. (b) Superposition of the three-dimensional crystal structures of P60_tth (violet) and B. cereus YkfC (grey; PDB entry 3h41). The strand and helix numbering corresponds to that of P60_tth. (c) Comparison of active sites in the two crystal structures displayed as a cross-eyed stereoview. The colour code is the same as in (b).
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fig3: Structure comparison of the catalytic domain of P60_tth with YkfC from B. cereus. (a) Primary-sequence alignment of P60_tth with YkfC from B. cereus. The secondary structures of the two proteins are indicated. The red sphere indicates the catalytic triad, while the blue spheres indicate other residues that are predicted to be involved in forming the catalytic site. (b) Superposition of the three-dimensional crystal structures of P60_tth (violet) and B. cereus YkfC (grey; PDB entry 3h41). The strand and helix numbering corresponds to that of P60_tth. (c) Comparison of active sites in the two crystal structures displayed as a cross-eyed stereoview. The colour code is the same as in (b).

Mentions: The catalytic domain is made up of a central β-sheet composed of five antiparallel β-strands that are surrounded by four α-helices (Figs. 1 ▶c and 3 ▶a). A search for structurally related proteins using the DALI server (Holm & Rosenström, 2010 ▶) shows that the catalytic domain matches structures from the NlpC/P60 protein family. The most similar structures are the putative cell-wall hydrolase from Clostridium difficile (PDB entry 4hpe; Joint Center for Structural Genomics, unpublished work), with a Z-score of 16.3 and a root-mean-square deviation (r.m.s.d.) of 2.7 Å over the Cα atoms of 116 residues, and the d,l-endopeptidase YkfC from B. cereus (PDB entry 3h41; Xu et al., 2010 ▶), with a Z-score of 16.1 and an r.m.s.d. of 1.9 Å over the Cα atoms of 111 residues. These two catalytic domains share 32% sequence identity with the catalytic domain of P60_tth. The comparison to the YkfC structure is of interest since it was solved with a bound ligand: the l-Ala-d-Glu peptide (Xu et al., 2010 ▶). As such, we can use the YkfC model to identify the putative catalytic residues of P60_tth and to propose its probable function.


An intermolecular binding mechanism involving multiple LysM domains mediates carbohydrate recognition by an endopeptidase.

Wong JE, Midtgaard SR, Gysel K, Thygesen MB, Sørensen KK, Jensen KJ, Stougaard J, Thirup S, Blaise M - Acta Crystallogr. D Biol. Crystallogr. (2015)

Structure comparison of the catalytic domain of P60_tth with YkfC from B. cereus. (a) Primary-sequence alignment of P60_tth with YkfC from B. cereus. The secondary structures of the two proteins are indicated. The red sphere indicates the catalytic triad, while the blue spheres indicate other residues that are predicted to be involved in forming the catalytic site. (b) Superposition of the three-dimensional crystal structures of P60_tth (violet) and B. cereus YkfC (grey; PDB entry 3h41). The strand and helix numbering corresponds to that of P60_tth. (c) Comparison of active sites in the two crystal structures displayed as a cross-eyed stereoview. The colour code is the same as in (b).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4356369&req=5

fig3: Structure comparison of the catalytic domain of P60_tth with YkfC from B. cereus. (a) Primary-sequence alignment of P60_tth with YkfC from B. cereus. The secondary structures of the two proteins are indicated. The red sphere indicates the catalytic triad, while the blue spheres indicate other residues that are predicted to be involved in forming the catalytic site. (b) Superposition of the three-dimensional crystal structures of P60_tth (violet) and B. cereus YkfC (grey; PDB entry 3h41). The strand and helix numbering corresponds to that of P60_tth. (c) Comparison of active sites in the two crystal structures displayed as a cross-eyed stereoview. The colour code is the same as in (b).
Mentions: The catalytic domain is made up of a central β-sheet composed of five antiparallel β-strands that are surrounded by four α-helices (Figs. 1 ▶c and 3 ▶a). A search for structurally related proteins using the DALI server (Holm & Rosenström, 2010 ▶) shows that the catalytic domain matches structures from the NlpC/P60 protein family. The most similar structures are the putative cell-wall hydrolase from Clostridium difficile (PDB entry 4hpe; Joint Center for Structural Genomics, unpublished work), with a Z-score of 16.3 and a root-mean-square deviation (r.m.s.d.) of 2.7 Å over the Cα atoms of 116 residues, and the d,l-endopeptidase YkfC from B. cereus (PDB entry 3h41; Xu et al., 2010 ▶), with a Z-score of 16.1 and an r.m.s.d. of 1.9 Å over the Cα atoms of 111 residues. These two catalytic domains share 32% sequence identity with the catalytic domain of P60_tth. The comparison to the YkfC structure is of interest since it was solved with a bound ligand: the l-Ala-d-Glu peptide (Xu et al., 2010 ▶). As such, we can use the YkfC model to identify the putative catalytic residues of P60_tth and to propose its probable function.

Bottom Line: The crystal structure and small-angle X-ray scattering solution studies of this endopeptidase revealed the presence of a homodimer.The structure of the two LysM domains co-crystallized with N-acetyl-chitohexaose revealed a new intermolecular binding mode that may explain the differential interaction between LysM domains and short or long chitin oligomers.By combining the structural information with the three-dimensional model of peptidoglycan, a model suggesting how protein dimerization enhances the recognition of peptidoglycan is proposed.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Carbohydrate Recognition and Signalling, Department of Molecular Biology and Genetics, Aarhus University, Gustav Wieds Vej 10C, 8000 Aarhus, Denmark.

ABSTRACT
LysM domains, which are frequently present as repetitive entities in both bacterial and plant proteins, are known to interact with carbohydrates containing N-acetylglucosamine (GlcNAc) moieties, such as chitin and peptidoglycan. In bacteria, the functional significance of the involvement of multiple LysM domains in substrate binding has so far lacked support from high-resolution structures of ligand-bound complexes. Here, a structural study of the Thermus thermophilus NlpC/P60 endopeptidase containing two LysM domains is presented. The crystal structure and small-angle X-ray scattering solution studies of this endopeptidase revealed the presence of a homodimer. The structure of the two LysM domains co-crystallized with N-acetyl-chitohexaose revealed a new intermolecular binding mode that may explain the differential interaction between LysM domains and short or long chitin oligomers. By combining the structural information with the three-dimensional model of peptidoglycan, a model suggesting how protein dimerization enhances the recognition of peptidoglycan is proposed.

Show MeSH
Related in: MedlinePlus