Mapping of the docking of SecA onto the chaperone SecB by site-directed spin labeling: insight into the mechanism of ligand transfer during protein export.
Export of protein into the periplasm of Escherichia coli via the general secretory system is achieved by action of a ternary complex comprising the polypeptide ligand, the chaperone SecB and SecA, a peripheral component of the membrane translocon, which is itself an ATPase. The unfolded ligand is captured initially by SecB and must be transferred to SecA and subsequently through the membrane translocon into the periplasm. We have taken the first steps in the elucidation of the mechanism of this dynamic transfer by determining the interface of interaction between SecB and SecA. Site-directed spin labeling and electron paramagnetic resonance spectroscopy were combined to identify which of the residues on SecB showed changes in spectral line shape upon addition of SecA. In all, 43% of the surface of SecB was covered by the 41 positions examined. A model of docking between SecB and SecA is proposed based on the pattern of amino acid residues on SecB shown to make contacts when in complex with SecA. This model in combination with previously published biochemical data provides insight into the transfer of the unfolded polypeptide from the chaperone SecB to SecA.
Crane
Jennine M
JM
Department of Biochemistry, University of Missouri, Columbia, MO 65211, USA.
Mao
Chunfeng
C
Lilly
Angela A
AA
Smith
Virginia F
VF
Suo
Yuying
Y
Hubbell
Wayne L
WL
Randall
Linda L
LL
D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D008024:Ligands; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D015394:Molecular Structure; D011485:Protein Binding; D011487:Protein Conformation; D021381:Protein Transport / Q000502:physiology*; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D013113:Spin Labels
D001426:Bacterial Proteins; D008024:Ligands; D026901:Membrane Transport Proteins; D000069816:SEC Translocation Channels; C057842:SecB protein, Bacteria; D013113:Spin Labels; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins