180-kD ribosome receptor is essential for both ribosome binding and protein translocation.
We have previously isolated a 180-kD ribosome receptor (p180) from mammalian rough ER that, when incorporated into liposomes, bound ribosomes with an affinity similar to intact membranes. To directly assess the contribution of p180 to ribosome binding as well as protein translocation, monoclonal antibodies were used to selectively deplete p180 from the detergent extracts of rough ER membranes used in the preparation of translocation-competent proteoliposomes. Proteoliposomes prepared from p180-depleted extracts showed a reduction in ribosome binding to the level of trypsin-inactivated controls as well as a loss in their ability to cotranslationally translocate two different secretory protein precursors. When purified p180 was added back to depleted extracts before proteoliposome formation, both ribosome binding and translocation activity were restored. In addition, the monoclonal antibodies, as well as their Fab' fragments, were able to inhibit ribosome binding and protein translocation when bound to intact rough microsomes. These data provide direct evidence that the 180-kD ribosome receptor is essential for ribosome binding and for the translocation of nascent proteins across the membrane of the rough ER.
Savitz
A J
AJ
Department of Biological Chemistry, University of California, Los Angeles School of Medicine.
Meyer
D I
DI
D000818:Animals; D001692:Biological Transport; D004285:Dogs; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000378:metabolism; D008970:Molecular Weight; D011506:Proteins / Q000378:metabolism*; D011510:Proteolipids; D011956:Receptors, Cell Surface / Q000276:immunology / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear; D012270:Ribosomes / Q000378:metabolism*
D008565:Membrane Proteins; D011506:Proteins; D011510:Proteolipids; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; C020485:proteoliposomes; C065526:ribosome receptor