Topology of the Salmonella invasion protein SipB in a model bilayer.
A critical early event in Salmonella infection is entry into intestinal epithelial cells. The Salmonella invasion protein SipB is required for the delivery of bacterial effector proteins into target eukaryotic cells, which subvert signal transduction pathways and cytoskeletal dynamics. SipB inserts into the host plasma membrane during infection, and the purified protein has membrane affinity and heterotypic membrane fusion activity in vitro. We used complementary biochemical and biophysical techniques to investigate the topology of purified SipB in a model membrane. We show that the 593 residue SipB is predominantly alpha-helical in aqueous solution, and that no significant change in secondary structural content accompanies lipid interaction. SipB contains two -helical transmembrane domains (residues 320-353 and 409-427), which insert deeply into the bilayer. Their integration allowed the hydrophilic region between the hydrophobic domains (354-408) to cross the bilayer. SipB membrane integration required both the hydrophobic domains and an additional helical C-terminal region (428-593). Further spectroscopic analysis of these domains in isolation showed that the hydrophobic regions insert obliquely into the bilayer, whereas the C-terminal domain associates with the bilayer surface, tilted parallel to the membrane. The combined data suggest a topological model for membrane-inserted SipB.
McGhie
Emma J
EJ
Department of Pathology, Cambridge University, Tennis Court Road, Cambridge CB2 1QP, UK.
Hume
Peter J
PJ
Hayward
Richard D
RD
Torres
Jaume
J
Koronakis
Vassilis
V
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002942:Circular Dichroism; D006801:Humans; D008027:Light; D008051:Lipid Bilayers / Q000378:metabolism*; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015335:Molecular Probes / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D015394:Molecular Structure; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012475:Salmonella / Q000378:metabolism*; D017550:Spectroscopy, Fourier Transform Infrared
D001426:Bacterial Proteins; D008051:Lipid Bilayers; D008081:Liposomes; D008565:Membrane Proteins; D015335:Molecular Probes; C094579:invasion protein B, Salmonella typhimurium