Development of a dna vaccine against _streptococcus mutans_ [electronic resource] : a novel approach to immunization against dental caries / Han, Thomas. [Tampa, Fla.] : University of South Florida, 2005. eng ABSTRACT: Streptococcus mutans is the main causative agent of dental caries, which is a widespread infectious disease. A number of surface molecules are involved in the pathogenicity of this organism, including adherence and aggregation factors. The wall-associated protein A (WapA) of Streptococcus mutans GS-5 was previously demonstrated to be a sucrose-dependent adherence and aggregation factor, and is a larger precursor to extracellular antigen A (AgA), a candidate antigen for a dental caries vaccine.The full-length wapA gene and a C-terminal truncated version agA encoding the AgA were cloned into the mammalian expression vector pcDNA 3.1/V5/His-TOPO. The above constructs were mixed with a cationic lipid and used to transfect Chinese hamster ovary (CHO) cells. Transient expression of the wapA and agA genes was observed at 24 h post-transfection, as shown by Western immunoblot analysis. In CHO, cells WapA containing the membrane and wall-spanning region was found in apoptotic bodies, whereas the soluble AgA, which lacked the hydrophobic region, was found in extracellular medium. A higher salivary IgA level was observed in mice immunized with the pcDNA-wapA vaccine as compared to those immunized with the pcDNA-agA vaccine. Furthermore, the anti-WapA antibody inhibited S. mutans sucrose-dependent adherence, suggesting potential protection of the tooth against S. mutans colonization, while anti-AgA had no significant effect. Indeed, prediction and analysis of protein epitopes showed that WapA contains highly promiscuous MHC-II binding motifs that are absent from AgA. Immunodot assay confirmed that WapA bound biotin-labeled dextran, whereas AgA did not. Thesis (Ph.D.)--University of South Florida, 2005. Includes bibliographical references. Text (Electronic thesis) in PDF format. System requirements: World Wide Web browser and PDF reader. Mode of access: World Wide Web. ABSTRACT: Streptococcus mutans is the main causative agent of dental caries, which is a widespread infectious disease. A number of surface molecules are involved in the pathogenicity of this organism, including adherence and aggregation factors. The wall-associated protein A (WapA) of Streptococcus mutans GS-5 was previously demonstrated to be a sucrose-dependent adherence and aggregation factor, and is a larger precursor to extracellular antigen A (AgA), a candidate antigen for a dental caries vaccine.The full-length wapA gene and a C-terminal truncated version agA encoding the AgA were cloned into the mammalian expression vector pcDNA 3.1/V5/His-TOPO. The above constructs were mixed with a cationic lipid and used to transfect Chinese hamster ovary (CHO) cells. Transient expression of the wapA and agA genes was observed at 24 h post-transfection, as shown by Western immunoblot analysis. In CHO, cells WapA containing the membrane and wall-spanning region was found in apoptotic bodies, whereas the soluble AgA, which lacked the hydrophobic region, was found in extracellular medium. A higher salivary IgA level was observed in mice immunized with the pcDNA-wapA vaccine as compared to those immunized with the pcDNA-agA vaccine. Furthermore, the anti-WapA antibody inhibited S. mutans sucrose-dependent adherence, suggesting potential protection of the tooth against S. mutans colonization, while anti-AgA had no significant effect. Indeed, prediction and analysis of protein epitopes showed that WapA contains highly promiscuous MHC-II binding motifs that are absent from AgA. Immunodot assay confirmed that WapA bound biotin-labeled dextran, whereas AgA did not. Adviser: My Lien Dao. Co-adviser: Mutans streptococcus. Prime-boost. Saliva. Secretory iga. Wapa.
Development of a dna vaccine against _streptococcus mutans_ [electronic resource] : a novel approach to immunization against dental caries /
Han, Thomas.
[Tampa, Fla.] : University of South Florida,
2005.
eng
ABSTRACT: Streptococcus mutans is the main causative agent of dental caries, which is a widespread infectious disease. A number of surface molecules are involved in the pathogenicity of this organism, including adherence and aggregation factors. The wall-associated protein A (WapA) of Streptococcus mutans GS-5 was previously demonstrated to be a sucrose-dependent adherence and aggregation factor, and is a larger precursor to extracellular antigen A (AgA), a candidate antigen for a dental caries vaccine.The full-length wapA gene and a C-terminal truncated version agA encoding the AgA were cloned into the mammalian expression vector pcDNA 3.1/V5/His-TOPO. The above constructs were mixed with a cationic lipid and used to transfect Chinese hamster ovary (CHO) cells. Transient expression of the wapA and agA genes was observed at 24 h post-transfection, as shown by Western immunoblot analysis.
In CHO, cells WapA containing the membrane and wall-spanning region was found in apoptotic bodies, whereas the soluble AgA, which lacked the hydrophobic region, was found in extracellular medium. A higher salivary IgA level was observed in mice immunized with the pcDNA-wapA vaccine as compared to those immunized with the pcDNA-agA vaccine. Furthermore, the anti-WapA antibody inhibited S. mutans sucrose-dependent adherence, suggesting potential protection of the tooth against S. mutans colonization, while anti-AgA had no significant effect. Indeed, prediction and analysis of protein epitopes showed that WapA contains highly promiscuous MHC-II binding motifs that are absent from AgA. Immunodot assay confirmed that WapA bound biotin-labeled dextran, whereas AgA did not.
Thesis (Ph.D.)--University of South Florida, 2005.
Includes bibliographical references.
Text (Electronic thesis) in PDF format.
System requirements: World Wide Web browser and PDF reader.
Mode of access: World Wide Web.
ABSTRACT: Streptococcus mutans is the main causative agent of dental caries, which is a widespread infectious disease. A number of surface molecules are involved in the pathogenicity of this organism, including adherence and aggregation factors. The wall-associated protein A (WapA) of Streptococcus mutans GS-5 was previously demonstrated to be a sucrose-dependent adherence and aggregation factor, and is a larger precursor to extracellular antigen A (AgA), a candidate antigen for a dental caries vaccine.The full-length wapA gene and a C-terminal truncated version agA encoding the AgA were cloned into the mammalian expression vector pcDNA 3.1/V5/His-TOPO. The above constructs were mixed with a cationic lipid and used to transfect Chinese hamster ovary (CHO) cells. Transient expression of the wapA and agA genes was observed at 24 h post-transfection, as shown by Western immunoblot analysis.
In CHO, cells WapA containing the membrane and wall-spanning region was found in apoptotic bodies, whereas the soluble AgA, which lacked the hydrophobic region, was found in extracellular medium. A higher salivary IgA level was observed in mice immunized with the pcDNA-wapA vaccine as compared to those immunized with the pcDNA-agA vaccine. Furthermore, the anti-WapA antibody inhibited S. mutans sucrose-dependent adherence, suggesting potential protection of the tooth against S. mutans colonization, while anti-AgA had no significant effect. Indeed, prediction and analysis of protein epitopes showed that WapA contains highly promiscuous MHC-II binding motifs that are absent from AgA. Immunodot assay confirmed that WapA bound biotin-labeled dextran, whereas AgA did not.
Adviser: My Lien Dao.
Co-adviser:
Mutans streptococcus.
Prime-boost.
Saliva.
Secretory iga.
Wapa.