Identification of the coiled-coil domains of Enterococcus faecalis DivIVA that mediate oligomerization and their importance for biological function.
Bacillus subtilis (Bs) DivIVA comprises coiled-coil structures and self-associates forming a 10-12 mer complex in vitro. Using bioinformatic approaches, we determined that Enterococcus faecalis (Ef) DivIVA comprises four coiled-coil domains, one at the N-terminus, the second and the third in the central region of the protein and the fourth at the C-terminus. We determined that DivIVA(Ef) self-interacts and forms a 10-12 multimeric complex. Point mutations or deletions of the central regions predicted bioinformatically to disrupt the coiled-coil structures either eliminated or weakened DivIVA(Ef) self-interaction and reduced oligomerization. Mutations disrupting the N- and C-terminal coiled-coils of DivIVA(Ef) did not affect DivIVA(Ef) oligomerization. The introduction of DivIVA(Ef) mutations to both the N-terminal and the central coiled-coil domains were lethal unless rescued by expressing wild-type DivIVA(Ef) in trans. E. faecalis cells expressing these mutations displayed aberrant cell morphology, indicating disruption of the normal cell division phenotype. The results in E. faecalis also indicate that both the N-terminal and the central coiled-coil structures of DivIVA(Ef) are indispensable for proper biological function. Overexpression of wild-type DivIVA(Ef) in both rod-shaped and round Escherichia coli cells resulted in morphological changes, while the overexpression of DivIVA(Ef) mutations failed to induce such alterations.
Rigden
Marc D
MD
Department of Biochemistry, Microbiology and Immunology, University of Ottawa, 451 Smyth Road, Ottawa, Ontario K1H 8M5, Canada.
Baier
Cherise
C
Ramirez-Arcos
Sandra
S
Liao
Mingmin
M
Wang
Monica
M
Dillon
Jo-Anne R
JA
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002455:Cell Division; D013293:Enterococcus faecalis / Q000166:cytology / Q000235:genetics; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics; D008970:Molecular Weight; D009154:Mutation; D017434:Protein Structure, Tertiary
D001426:Bacterial Proteins; D018797:Cell Cycle Proteins; C104636:DivIVA protein, bacteria