Expression of Streptococcus mutans wall-associated protein A gene in Chinese hamster ovary cells: prospect for a dental caries DNA vaccine.
The Streptococcus mutans strain GS-5 wall-associated protein A (Wap-A) is a precursor to the extracellular antigen A (AgA), a recognized candidate dental caries vaccine. The full-length wapA gene (wapA-E) and a C-terminal truncated version (wapA-G) encoding the AgA were cloned into the mammalian expression vector pcDNA 3.1/V5/His-TOPO. The resulting constructs were propagated in the Escherichia coli Top10. To investigate the expression of the S. mutans genes in mammalian cells, the above constructs were used to transfect Chinese hamster ovary (CHO) cells in the presence of the cationic lipid pfx-8. Transient expression of the wapA-E and wapA-G genes was observed at 24 h post-transfection, as shown by Western immunoblot analysis using a rabbit antiserum to S. mutans cell wall. Immunochemical staining of the transfected CHO cells showed expression of WapA mainly in the cells and budding vesicles, whereas AgA was found mainly in the transfected cells and extracellular medium. The expression of S. mutans proteins in CHO cells, in either vesicles or soluble form, suggested an antibody response to the above DNA constructs. Work is under way to test the efficacy of these as DNA vaccines against S. mutans.
Han
T K
TK
Department of Biology, College of Arts and Sciences, University of South Florida, Tampa, Florida 33620-5150, USA.
Yoder
S
S
Cao
C
C
Ugen
K E
KE
Dao
M L
ML
D000818:Animals; D000942:Antigens, Bacterial; D001426:Bacterial Proteins / Q000235:genetics*; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D003731:Dental Caries / Q000382:microbiology / Q000517:prevention & control*; D011817:Rabbits; D022541:Streptococcal Vaccines / Q000235:genetics*; D013295:Streptococcus mutans / Q000235:genetics* / Q000276:immunology*; D014162:Transfection; D019444:Vaccines, DNA / Q000235:genetics*
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D022541:Streptococcal Vaccines; D019444:Vaccines, DNA; C081813:wapA protein, Bacillus subtilis