Role of the C terminus in antigen P1 surface localization in Streptococcus mutans and two related cocci.
The C terminus of the major surface protein P1 from Streptococcus mutans is composed of a hydrophilic domain, an LPNTGV motif, a hydrophobic domain, and a charged tail. These features are shared by surface proteins from many gram-positive coccal bacteria. To investigate the role of the C-terminal domains in antigen P1 surface localization, full-length and truncated P1 gene constructs, which were expressed on the shuttle vector pDL276, were transformed into the P1-negative mutant S. mutans SM3352, Streptococcus gordonii DL-1, and Enterococcus faecalis UV202. Transformants were tested for expression of P1 by enzyme-linked immunosorbent assaying and Western blotting. The results showed that full-length P1 was expressed by transformants of all three bacteria and was localized on the cell surface. A fusion protein composed of the Staphylococcus aureus fibronectin binding protein C terminus and the P1 protein N terminus was found to surface localize in S. mutans. Deletion of the entire C-terminal domains resulted in P1 being expressed in the culture supernatant. A P1 truncation, which carried only the hydrophilic domain at its C terminus, was found partially associated with the cell surface. This truncated P1 was readily removed from the isolated cell wall by hot sodium dodecyl sulfate-mercaptoethanol extraction. In contrast, the full-length P1 remained associated with the isolated cell wall after similar treatment, suggesting covalent linkages between the full-length P1 and the cell wall. The results described above showed that antigen P1 was anchored to the cell wall by its C-terminal domains probably via covalent linkages with the cell wall. The results also support a universal mechanism involving the C-terminal domains for protein surface localization among this group of gram-positive bacteria.
Department of Oral Biology, Faculty of Dentistry, University of Manitoba, Winnipeg, Canada.
M K
MK
Homonylo-McGavin
S F
SF
Lee
D000595:Amino Acid Sequence; D000818:Animals; D000942:Antigens, Bacterial / Q000032:analysis* / Q000737:chemistry; D001426:Bacterial Proteins / Q000032:analysis* / Q000737:chemistry; D001483:Base Sequence; D002473:Cell Wall / Q000737:chemistry; D013293:Enterococcus faecalis / Q000276:immunology*; D005455:Fluorescent Antibody Technique; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data; D009154:Mutation; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D013291:Streptococcus / Q000276:immunology*; D013295:Streptococcus mutans / Q000276:immunology*
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; C055764:S-layer proteins