The identification of proteins in the proximity of signal-anchor sequences during their targeting to and insertion into the membrane of the ER.
Using a photocross-linking approach we have investigated the cytosolic and membrane components involved in the targeting and insertion of signal-anchor proteins into the membrane of the ER. The nascent chains of both type I and type II signal-anchor proteins can be cross-linked to the 54-kD subunit of the signal recognition particle. Upon addition of rough microsomes the type I and type II signal-anchor proteins interact with a number of components. Both types of protein interact with an integral membrane protein, the signal sequence receptor, previously identified by its proximity to preprolactin during its translocation (Wiedmann, M., T.V. Kurzchalia, E. Hartmann, and T.A. Rapoport. 1987. Nature [Lond.] 328:830-833). Three proteins, previously unidentified, were found to be cross-linked to the nascent chains of the signal-anchor proteins. Among them was a 37-kD protein that was found to be the main component interacting with the type I SA protein used. These proteins were not seen in the absence of membranes suggesting they are components of the ER. The ability of the nascent chains to be cross-linked to these identified proteins was shown to be abolished by prior treatment with agents known to disrupt translocation intermediates or ribosomes. We propose that the newly identified proteins function either in the membrane insertion of only a subset of proteins or only at a specific stage of insertion.
High
S
S
European Molecular Biology Laboratory, Heidelberg, Germany.
Görlich
D
D
Wiedmann
M
M
Rapoport
T A
TA
Dobberstein
B
B
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D002451:Cell Compartmentation; D003432:Cross-Linking Reagents; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011498:Protein Precursors / Q000378:metabolism; D021382:Protein Sorting Signals / Q000502:physiology*; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012261:Ribonucleoproteins / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism; D018271:Signal Recognition Particle; D014357:Trypsin / Q000494:pharmacology
D003432:Cross-Linking Reagents; D008565:Membrane Proteins; D011498:Protein Precursors; D021382:Protein Sorting Signals; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; D018271:Signal Recognition Particle; D014357:Trypsin