Molecular Insights on the Recognition of a Lactococcus lactis Cell Wall Pellicle by the Phage 1358 Receptor Binding Protein

ABSTRACT The Gram-positive bacterium Lactococcus lactis is used for the production of cheeses and other fermented dairy products. Accidental infection of L. lactis cells by virulent lactococcal tailed phages is one of the major risks of fermentation failures in industrial dairy factories. Lactococcal phage 1358 possesses a host range limited to a few L. lactis strains and strong genomic similarities to Listeria phages. We report here the X-ray structures of phage 1358 receptor binding protein (RBP) in complex with monosaccharides. Each monomer of its trimeric RBP is formed of two domains: a “shoulder” domain linking the RBP to the rest of the phage and a jelly roll fold “head/host recognition” domain. This domain harbors a saccharide binding crevice located in the middle of a monomer. Crystal structures identified two sites at the RBP surface, ∼8 Å from each other, one accommodating a GlcNAc monosaccharide and the other accommodating a GlcNAc or a glucose 1-phosphate (Glc1P) monosaccharide. GlcNAc and GlcNAc1P are components of the polysaccharide pellicle that we identified at the cell surface of L. lactis SMQ-388, the host of phage 1358. We therefore modeled a galactofuranose (Galf) sugar bridging the two GlcNAc saccharides, suggesting that the trisaccharidic motif GlcNAc-Galf-GlcNAc (or Glc1P) might be common to receptors of genetically distinct lactococcal phages p2, TP091-1, and 1358. Strain specificity might therefore be elicited by steric clashes induced by the remaining components of the pellicle hexasaccharide. Taken together, these results provide a first insight into the molecular mechanism of host receptor recognition by lactococcal phages. IMPORTANCE Siphophages infecting the Gram-positive bacterium Lactococcus lactis are sources of milk fermentation failures in the dairy industry. We report here the structure of the pellicle polysaccharide from L. lactis SMQ-388, the specific host strain of phage 1358. We determined the X-ray structures of the lytic lactococcal phage 1358 receptor binding protein (RBP) in complex with monosaccharides. The positions and nature of monosaccharides bound to the RBP are in agreement with the pellicle structure and suggest a general binding mode of lactococcal phages to their pellicle saccharidic receptor.

[1]  C. Cambillau,et al.  Differences in Lactococcal Cell Wall Polysaccharide Structure Are Major Determining Factors in Bacteriophage Sensitivity , 2014, mBio.

[2]  S. Lortal,et al.  Structural studies of the cell wall polysaccharides from three strains of Lactobacillus helveticus with different autolytic properties: DPC4571, BROI, and LH1. , 2013, Carbohydrate research.

[3]  H. Neve,et al.  Investigation of the Relationship between Lactococcal Host Cell Wall Polysaccharide Genotype and 936 Phage Receptor Binding Protein Phylogeny , 2013, Applied and Environmental Microbiology.

[4]  H. Neve,et al.  Identification of a New P335 Subgroup through Molecular Analysis of Lactococcal Phages Q33 and BM13 , 2013, Applied and Environmental Microbiology.

[5]  A. Desmyter,et al.  Viral infection modulation and neutralization by camelid nanobodies , 2013, Proceedings of the National Academy of Sciences.

[6]  M. van Heel,et al.  Visualizing a Complete Siphoviridae Member by Single-Particle Electron Microscopy: the Structure of Lactococcal Phage TP901-1 , 2012, Journal of Virology.

[7]  M. Paternostre,et al.  New insights into pb5, the receptor binding protein of bacteriophage T5, and its interaction with its Escherichia coli receptor FhuA. , 2012, Biochimie.

[8]  D. Veesler,et al.  Structure of the phage TP901-1 1.8 MDa baseplate suggests an alternative host adhesion mechanism , 2012, Proceedings of the National Academy of Sciences.

[9]  S. Moineau,et al.  Bacteriophages of lactic acid bacteria and their impact on milk fermentations , 2011, Microbial cell factories.

[10]  D. Veesler,et al.  Unraveling Lactococcal Phage Baseplate Assembly by Mass Spectrometry , 2011, Molecular & Cellular Proteomics.

[11]  M. van Heel,et al.  Structure and Molecular Assignment of Lactococcal Phage TP901-1 Baseplate* , 2010, The Journal of Biological Chemistry.

[12]  Guillaume Andre,et al.  Imaging the nanoscale organization of peptidoglycan in living Lactococcus lactis cells , 2010, Nature communications.

[13]  G. Sciara,et al.  Structure of lactococcal phage p2 baseplate and its mechanism of activation , 2010, Proceedings of the National Academy of Sciences.

[14]  Kevin Cowtan,et al.  Recent developments in classical density modification , 2010, Acta crystallographica. Section D, Biological crystallography.

[15]  P. Emsley,et al.  Features and development of Coot , 2010, Acta crystallographica. Section D, Biological crystallography.

[16]  C. Péchoux,et al.  Cell Surface of Lactococcus lactis Is Covered by a Protective Polysaccharide Pellicle* , 2010, The Journal of Biological Chemistry.

[17]  S. Moineau,et al.  Genome Organization and Characterization of the Virulent Lactococcal Phage 1358 and Its Similarities to Listeria Phages , 2010, Applied and Environmental Microbiology.

[18]  M. Loessner,et al.  Comparative Genome Analysis of Listeria Bacteriophages Reveals Extensive Mosaicism, Programmed Translational Frameshifting, and a Novel Prophage Insertion Site , 2009, Journal of bacteriology.

[19]  Andreas Plückthun,et al.  Crystal Structure and Function of a DARPin Neutralizing Inhibitor of Lactococcal Phage TP901-1 , 2009, The Journal of Biological Chemistry.

[20]  C. Cambillau,et al.  Moineau, Christian Cambillau and Valérie Campanacci Lactococcal Phages Binding Protein Constructed from Two Crystal Structure of a Chimeric Receptor Supplemental Material , 2008 .

[21]  G. Sciara,et al.  A Topological Model of the Baseplate of Lactococcal Phage Tuc2009* , 2008, Journal of Biological Chemistry.

[22]  L. Hammarström,et al.  Lactobacillli expressing llama VHH fragments neutralise Lactococcus phages , 2007, BMC biotechnology.

[23]  Randy J. Read,et al.  Phaser crystallographic software , 2007, Journal of applied crystallography.

[24]  C. Cambillau,et al.  Crystal Structure of the Receptor-Binding Protein Head Domain from Lactococcus lactis Phage bIL170 , 2006, Journal of Virology.

[25]  Kevin Cowtan,et al.  The Buccaneer software for automated model building. 1. Tracing protein chains. , 2006, Acta crystallographica. Section D, Biological crystallography.

[26]  Kevin Cowtan,et al.  The Buccaneer software for automated model building , 2006 .

[27]  Liang Tang,et al.  The Structure of an Infectious P22 Virion Shows the Signal for Headful DNA Packaging , 2006, Science.

[28]  H. Deveau,et al.  Biodiversity and Classification of Lactococcal Phages , 2006, Applied and Environmental Microbiology.

[29]  Sylvain Moineau,et al.  Modular Structure of the Receptor Binding Proteins of Lactococcus lactis Phages , 2006, Journal of Biological Chemistry.

[30]  A. Desmyter,et al.  Receptor-Binding Protein of Lactococcus lactis Phages: Identification and Characterization of the Saccharide Receptor-Binding Site , 2006, Journal of bacteriology.

[31]  R. Benz,et al.  Interaction of bacteriophage lambda with its cell surface receptor: an in vitro study of binding of the viral tail protein gpJ to LamB (Maltoporin). , 2006, Biochemistry.

[32]  R. Vincentelli,et al.  Automated expression and solubility screening of His-tagged proteins in 96-well format. , 2005, Analytical biochemistry.

[33]  Fumio Arisaka,et al.  The tail structure of bacteriophage T4 and its mechanism of contraction , 2005, Nature Structural &Molecular Biology.

[34]  Sylvain Moineau,et al.  Llama Antibodies against a Lactococcal Protein Located at the Tip of the Phage Tail Prevent Phage Infection , 2005, Journal of bacteriology.

[35]  Johannes Söding,et al.  The HHpred interactive server for protein homology detection and structure prediction , 2005, Nucleic Acids Res..

[36]  G. Bricogne,et al.  Refinement of severely incomplete structures with maximum likelihood in BUSTER-TNT. , 2004, Acta crystallographica. Section D, Biological crystallography.

[37]  Renaud Vincentelli,et al.  Medium-scale structural genomics: strategies for protein expression and crystallization. , 2003, Accounts of chemical research.

[38]  George M Sheldrick,et al.  Substructure solution with SHELXD. , 2002, Acta crystallographica. Section D, Biological crystallography.

[39]  N. Sharon,et al.  How proteins bind carbohydrates: lessons from legume lectins. , 2002, Journal of agricultural and food chemistry.

[40]  G. Murshudov,et al.  Refinement of macromolecular structures by the maximum-likelihood method. , 1997, Acta crystallographica. Section D, Biological crystallography.

[41]  C. Cambillau,et al.  Structures of a legume lectin complexed with the human lactotransferrin N2 fragment, and with an isolated biantennary glycopeptide: role of the fucose moiety. , 1994, Structure.

[42]  W. Sandine,et al.  A membrane protein is required for bacteriophage c2 infection of Lactococcus lactis subsp. lactis C2 , 1991, Journal of bacteriology.

[43]  I. Katsura.  Determination of bacteriophage λ tail length by a protein ruler , 1987, Nature.

[44]  J. Defaye,et al.  Conformations in Solution of α,α-Trehalose, α-d-Glucopyranosyl α-d-Mannopyranoside, and Their 1-Thioglycosyl Analogs, and a Tentative Correlation of Their Behaviour with Respect to the Enzyme Trehalase , 1983 .

[45]  H. Ackermann.  Bacteriophage electron microscopy. , 2012, Advances in virus research.

[46]  Alexei Vagin,et al.  Molecular replacement with MOLREP. , 2010, Acta crystallographica. Section D, Biological crystallography.

[47]  A. Plückthun,et al.  Crystal structure and function of a DARPin neutralizing inhibitor of lactococcal phage TP901-1: comparison of DARPin and camelid VHH binding mode , 2010 .

[48]  Sylvain Moineau,et al.  Lactococcal bacteriophage p2 receptor-binding protein structure suggests a common ancestor gene with bacterial and mammalian viruses , 2006, Nature Structural &Molecular Biology.

[49]  W. Sandine,et al.  Lactococcus lactis ssp. lactis C2 Bacteriophage sk1 Receptor involving Rhamnose and Glucose Moieties in the Cell Wall , 1994 .

[50]  I. Katsura.  Determination of bacteriophage lambda tail length by a protein ruler. , 1987, Nature.