Previous grant support has led to the discovery of a novel Streptococcus parasanguis fimbriae associated adhesion, Fapl. Fapl-like molecules and genes involved in Fapl glycosylation are strikingly conserved throughout oral streptococci and are also found in important pathogens, such as Streptococcus pneumoniae and Staphylococcus aureus. We will use S. parasanguis as a model system to study the function of Fapl-like proteins in these pathogenic bacteria. Fapl is required for the formation of a S. parasanguis biofilm. We have determined that the N-terminal polypeptide of Fapl (rFapl) is essential for initial attachment of bacteria to the biofilm surface. Fapl is glycosylated, and glycosylation of this protein is involved in maturation of biofilm. The overall goal of this application is highlighted by two Specific Aims: (1) to determine the structure/function relationship of the rFapl adhesin that is required for initial bacterial attachment and (2) to define the functions of the fapl flanking genes in Fapl glycosylation and bacterial biofilm formation. We will bring genetic, structural biology (NMR spectroscopy and X-ray crystallography), in vitro binding biochemical assays and in vivo animal model studies together to address how the structure of the Fapl polypeptide influences its function. These comprehensive approaches should shed light on the structural basis for binding of Fapl to salivary receptors, and for predicted Fapl-like molecules of other medically important pathogens. Proteins of the fapl flanking region may constitute a coupled secretion and glycosylation pathway for Fapl. To begin to understand the mechanisms involved, we will determine the subcellular Iocalization of proteins in this locus and identify putative protein-protein interactions in vivo and in vitro. We previously demonstrated that Fapl glycosylation is involved in maturation of the biofilm. Using a combination of molecular and genetic analyses, we will dissect the function of these glycosylation genes in biofilm formation. Elucidation of the function of Fapl glycosylation and definition of the Fapl polypeptide structure/function relationships will contribute to the field of bacterial colonization in the oral environment and to the pathogenic mechanism of other important pathogens as their genomes indicate that they also possess Fapl-like molecules, and the flanking proteins.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
2R01DE011000-11
Application #
6724676
Study Section
Oral Biology and Medicine Subcommittee 1 (OBM)
Program Officer
Mangan, Dennis F
Project Start
1994-05-01
Project End
2009-02-28
Budget Start
2004-06-01
Budget End
2005-02-28
Support Year
11
Fiscal Year
2004
Total Cost
$466,916
Indirect Cost
Name
University of Vermont & St Agric College
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
066811191
City
Burlington
State
VT
Country
United States
Zip Code
05405
Zhu, Fan; Erlandsen, Heidi; Ding, Lei et al. (2011) Structural and functional analysis of a new subfamily of glycosyltransferases required for glycosylation of serine-rich streptococcal adhesins. J Biol Chem 286:27048-57
Ramboarina, Stéphanie; Garnett, James A; Zhou, Meixian et al. (2010) Structural insights into serine-rich fimbriae from Gram-positive bacteria. J Biol Chem 285:32446-57
Zhou, Meixian; Zhu, Fan; Dong, Shengli et al. (2010) A novel glucosyltransferase is required for glycosylation of a serine-rich adhesin and biofilm formation by Streptococcus parasanguinis. J Biol Chem 285:12140-8
Zhou, Meixian; Wu, Hui (2009) Glycosylation and biogenesis of a family of serine-rich bacterial adhesins. Microbiology 155:317-27
Li, Yirong; Chen, Yabing; Huang, Xiang et al. (2008) A conserved domain of previously unknown function in Gap1 mediates protein-protein interaction and is required for biogenesis of a serine-rich streptococcal adhesin. Mol Microbiol 70:1094-104
Peng, Zhixiang; Fives-Taylor, Paula; Ruiz, Teresa et al. (2008) Identification of critical residues in Gap3 of Streptococcus parasanguinis involved in Fap1 glycosylation, fimbrial formation and in vitro adhesion. BMC Microbiol 8:52
Bu, Su; Li, Yirong; Zhou, Meixian et al. (2008) Interaction between two putative glycosyltransferases is required for glycosylation of a serine-rich streptococcal adhesin. J Bacteriol 190:1256-66
Peng, Z; Wu, H; Ruiz, T et al. (2008) Role of gap3 in Fap1 glycosylation, stability, in vitro adhesion, and fimbrial and biofilm formation of Streptococcus parasanguinis. Oral Microbiol Immunol 23:70-8
Zhou, Meixian; Peng, Zhixiang; Fives-Taylor, Paula et al. (2008) A conserved C-terminal 13-amino-acid motif of Gap1 is required for Gap1 function and necessary for the biogenesis of a serine-rich glycoprotein of Streptococcus parasanguinis. Infect Immun 76:5624-31
Zhou, Meixian; Fives-Taylor, Paula; Wu, Hui (2008) The utility of affinity-tags for detection of a streptococcal protein from a variety of streptococcal species. J Microbiol Methods 72:249-56

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