Interactions of the type III secretion pathway proteins LcrV and LcrG from Yersinia pestis are mediated by coiled-coil domains.
The type III secretion system is used by pathogenic Yersinia to translocate virulence factors into the host cell. A key component is the multifunctional LcrV protein, which is present on the bacterial surface prior to host cell contact and up-regulates translocation by blocking the repressive action of the LcrG protein on the cytosolic side of the secretion apparatus. The functions of LcrV are proposed to involve self-interactions (multimerization) and interactions with other proteins including LcrG. Coiled-coil motifs predicted to be present are thought to play a role in mediating these protein-protein interactions. We have purified recombinant LcrV, LcrG, and site-directed mutants of LcrV and demonstrated the structural integrity of these proteins using circular dichroism spectroscopy. We show that LcrV interacts both with itself and with LcrG and have obtained micromolar and nanomolar affinities for these interactions, respectively. The effects of LcrV mutations upon LcrG binding suggest that coiled-coil interactions indeed play a significant role in complex formation. In addition, comparisons of secretion patterns of effector proteins in Yersinia, arising from wild type and mutants of LcrV, support the proposed role of LcrG in titration of LcrV in vivo but also suggest that other factors may be involved.
Lawton
Daniel G
DG
Department of Biological Sciences, Centre for Molecular Microbiology and Infection, Flowers Building, Imperial College of Science, Technology and Medicine, London SW7 2AY, United Kingdom.
Longstaff
Colin
C
Wallace
B A
BA
Hill
Jim
J
Leary
Sophie E C
SE
Titball
Richard W
RW
Brown
Katherine A
KA
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012634:Bodily Secretions / Q000502:physiology*; D002942:Circular Dichroism; D008954:Models, Biological; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D052899:Pore Forming Cytotoxic Proteins; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020349:Surface Plasmon Resonance; D015010:Yersinia pestis / Q000378:metabolism*
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C088009:LcrG protein, Yersinia; C043908:LcrV protein, Yersinia; D052899:Pore Forming Cytotoxic Proteins; D011993:Recombinant Fusion Proteins