Signal sequence recognition in cotranslational translocation by protein components of the endoplasmic reticulum membrane.
We have investigated the role of membrane proteins and lipids during early phases of the cotranslational insertion of secretory proteins into the translocation channel of the endoplasmic reticulum (ER) membrane. We demonstrate that all steps, including the one during which signal sequence recognition occurs, can be reproduced with purified translocation components in detergent solution, in the absence of bulk lipids or a bilayer. Photocross-linking experiments with native membranes show that upon complete insertion into the channel signal sequences are both precisely positioned with respect to the protein components of the channel and contact lipids. Together, these results indicate that signal sequences are bound to a specific binding site at the interface between the channel and the surrounding lipids, and are recognized ultimately by protein-protein interactions. Our data also suggest that at least some signal sequences reach the binding site by transfer through the interior of the channel.
Mothes
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W
Howard Hughes Medical Institute and Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA.
Jungnickel
B
B
Brunner
J
J
Rapoport
T A
TA
D000818:Animals; D001665:Binding Sites; D001693:Biological Transport, Active; D003432:Cross-Linking Reagents; D003902:Detergents; D004285:Dogs; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D008563:Membrane Lipids / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D011388:Prolactin / Q000235:genetics / Q000378:metabolism; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism; D000069816:SEC Translocation Channels; D029701:Saccharomyces cerevisiae Proteins; D018271:Signal Recognition Particle / Q000235:genetics / Q000378:metabolism*; D012996:Solutions
D003432:Cross-Linking Reagents; D003902:Detergents; D005656:Fungal Proteins; C048632:MF(ALPHA)1 protein, S cerevisiae; D008563:Membrane Lipids; D008565:Membrane Proteins; D011498:Protein Precursors; D000069816:SEC Translocation Channels; D029701:Saccharomyces cerevisiae Proteins; D018271:Signal Recognition Particle; D012996:Solutions; C017671:preprolactin; D011388:Prolactin