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Structural determinants for activity and specificity of the bacterial toxin LlpA.

Ghequire MG, Garcia-Pino A, Lebbe EK, Spaepen S, Loris R, De Mot R - PLoS Pathog. (2013)

Bottom Line: The N-terminal MMBL domain (N-domain) adopts the same fold but is structurally more divergent and lacks a functional mannose-binding site.Differential activity of engineered N/C-domain chimers derived from two LlpA homologues with different killing spectra, disclosed that the N-domain determines target specificity.Apparently this bacteriocin is assembled from two structurally similar domains that evolved separately towards dedicated functions in target recognition and bacteriotoxicity.

View Article: PubMed Central - PubMed

Affiliation: Centre of Microbial and Plant Genetics, University of Leuven, Heverlee-Leuven, Belgium.

ABSTRACT
Lectin-like bacteriotoxic proteins, identified in several plant-associated bacteria, are able to selectively kill closely related species, including several phytopathogens, such as Pseudomonas syringae and Xanthomonas species, but so far their mode of action remains unrevealed. The crystal structure of LlpABW, the prototype lectin-like bacteriocin from Pseudomonas putida, reveals an architecture of two monocot mannose-binding lectin (MMBL) domains and a C-terminal β-hairpin extension. The C-terminal MMBL domain (C-domain) adopts a fold very similar to MMBL domains from plant lectins and contains a binding site for mannose and oligomannosides. Mutational analysis indicates that an intact sugar-binding pocket in this domain is crucial for bactericidal activity. The N-terminal MMBL domain (N-domain) adopts the same fold but is structurally more divergent and lacks a functional mannose-binding site. Differential activity of engineered N/C-domain chimers derived from two LlpA homologues with different killing spectra, disclosed that the N-domain determines target specificity. Apparently this bacteriocin is assembled from two structurally similar domains that evolved separately towards dedicated functions in target recognition and bacteriotoxicity.

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Overall structure of LlpABW.(A) Topology diagram of LlpABW. The N-domain is shown in red, the C-domain in blue and the C-terminal extension in green. The different strands and subdomains are labeled. Domain swapping involves β-strand segments β11b and β22b, which together with β-strand segments β11a and β22a link both MMBL domains. (B) Cartoon representation of LlpABW with the different domains colored as in panel A. The bound Me-Man residue is shown as an orange stick representation. (C) Domain orientations of LlpABW compared with the heterodimeric MMBL ASA I (Allium sativum agglutinin, PDB entry 1KJ1) and tandem MMBL SCAfet (Scilla campanulata fetuin-binding lectin, PDB entry 1DLP). In each case, the C-domain is shown in the same orientation, highlighting the different relative orientation of the N-domain in LlpABW. Domain-swapped dimers in homo-oligomeric plant MMBL lectins such as snowdrop lectin have their domain orientation similar to ASA I and SCAfet.
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ppat-1003199-g001: Overall structure of LlpABW.(A) Topology diagram of LlpABW. The N-domain is shown in red, the C-domain in blue and the C-terminal extension in green. The different strands and subdomains are labeled. Domain swapping involves β-strand segments β11b and β22b, which together with β-strand segments β11a and β22a link both MMBL domains. (B) Cartoon representation of LlpABW with the different domains colored as in panel A. The bound Me-Man residue is shown as an orange stick representation. (C) Domain orientations of LlpABW compared with the heterodimeric MMBL ASA I (Allium sativum agglutinin, PDB entry 1KJ1) and tandem MMBL SCAfet (Scilla campanulata fetuin-binding lectin, PDB entry 1DLP). In each case, the C-domain is shown in the same orientation, highlighting the different relative orientation of the N-domain in LlpABW. Domain-swapped dimers in homo-oligomeric plant MMBL lectins such as snowdrop lectin have their domain orientation similar to ASA I and SCAfet.

Mentions: The crystal structure of LlpABW from P. putida BW11M1 (LlpABW) shows it contains two β-prism MMBL domains, referred to as the N-domain and the C-domain following their position in the amino acid sequence (Figure 1A,B; Figure S1). The N-domain spans residues Arg4-Pro135 while the C-domain encompasses residues Ala136-Gln253. Each domain exhibits pseudo-threefold symmetry and the corresponding subdomains will be referred to as IN, IIN, IIIN, IC, IIC and IIIC, respectively (Figure 1A and Figure S1). Following these two domains, a β-hairpin extension is formed by residues Pro254-His275 (the numbering used in this paper corresponds to that of the wild-type protein without His-tag [21]).


Structural determinants for activity and specificity of the bacterial toxin LlpA.

Ghequire MG, Garcia-Pino A, Lebbe EK, Spaepen S, Loris R, De Mot R - PLoS Pathog. (2013)

Overall structure of LlpABW.(A) Topology diagram of LlpABW. The N-domain is shown in red, the C-domain in blue and the C-terminal extension in green. The different strands and subdomains are labeled. Domain swapping involves β-strand segments β11b and β22b, which together with β-strand segments β11a and β22a link both MMBL domains. (B) Cartoon representation of LlpABW with the different domains colored as in panel A. The bound Me-Man residue is shown as an orange stick representation. (C) Domain orientations of LlpABW compared with the heterodimeric MMBL ASA I (Allium sativum agglutinin, PDB entry 1KJ1) and tandem MMBL SCAfet (Scilla campanulata fetuin-binding lectin, PDB entry 1DLP). In each case, the C-domain is shown in the same orientation, highlighting the different relative orientation of the N-domain in LlpABW. Domain-swapped dimers in homo-oligomeric plant MMBL lectins such as snowdrop lectin have their domain orientation similar to ASA I and SCAfet.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585409&req=5

ppat-1003199-g001: Overall structure of LlpABW.(A) Topology diagram of LlpABW. The N-domain is shown in red, the C-domain in blue and the C-terminal extension in green. The different strands and subdomains are labeled. Domain swapping involves β-strand segments β11b and β22b, which together with β-strand segments β11a and β22a link both MMBL domains. (B) Cartoon representation of LlpABW with the different domains colored as in panel A. The bound Me-Man residue is shown as an orange stick representation. (C) Domain orientations of LlpABW compared with the heterodimeric MMBL ASA I (Allium sativum agglutinin, PDB entry 1KJ1) and tandem MMBL SCAfet (Scilla campanulata fetuin-binding lectin, PDB entry 1DLP). In each case, the C-domain is shown in the same orientation, highlighting the different relative orientation of the N-domain in LlpABW. Domain-swapped dimers in homo-oligomeric plant MMBL lectins such as snowdrop lectin have their domain orientation similar to ASA I and SCAfet.
Mentions: The crystal structure of LlpABW from P. putida BW11M1 (LlpABW) shows it contains two β-prism MMBL domains, referred to as the N-domain and the C-domain following their position in the amino acid sequence (Figure 1A,B; Figure S1). The N-domain spans residues Arg4-Pro135 while the C-domain encompasses residues Ala136-Gln253. Each domain exhibits pseudo-threefold symmetry and the corresponding subdomains will be referred to as IN, IIN, IIIN, IC, IIC and IIIC, respectively (Figure 1A and Figure S1). Following these two domains, a β-hairpin extension is formed by residues Pro254-His275 (the numbering used in this paper corresponds to that of the wild-type protein without His-tag [21]).

Bottom Line: The N-terminal MMBL domain (N-domain) adopts the same fold but is structurally more divergent and lacks a functional mannose-binding site.Differential activity of engineered N/C-domain chimers derived from two LlpA homologues with different killing spectra, disclosed that the N-domain determines target specificity.Apparently this bacteriocin is assembled from two structurally similar domains that evolved separately towards dedicated functions in target recognition and bacteriotoxicity.

View Article: PubMed Central - PubMed

Affiliation: Centre of Microbial and Plant Genetics, University of Leuven, Heverlee-Leuven, Belgium.

ABSTRACT
Lectin-like bacteriotoxic proteins, identified in several plant-associated bacteria, are able to selectively kill closely related species, including several phytopathogens, such as Pseudomonas syringae and Xanthomonas species, but so far their mode of action remains unrevealed. The crystal structure of LlpABW, the prototype lectin-like bacteriocin from Pseudomonas putida, reveals an architecture of two monocot mannose-binding lectin (MMBL) domains and a C-terminal β-hairpin extension. The C-terminal MMBL domain (C-domain) adopts a fold very similar to MMBL domains from plant lectins and contains a binding site for mannose and oligomannosides. Mutational analysis indicates that an intact sugar-binding pocket in this domain is crucial for bactericidal activity. The N-terminal MMBL domain (N-domain) adopts the same fold but is structurally more divergent and lacks a functional mannose-binding site. Differential activity of engineered N/C-domain chimers derived from two LlpA homologues with different killing spectra, disclosed that the N-domain determines target specificity. Apparently this bacteriocin is assembled from two structurally similar domains that evolved separately towards dedicated functions in target recognition and bacteriotoxicity.

Show MeSH
Related in: MedlinePlus