GTPase domain of the 54-kD subunit of the mammalian signal recognition particle is required for protein translocation but not for signal sequence binding.
The 54-kD subunit of the signal recognition particle (SRP54) binds to signal sequences of nascent secretory and transmembrane proteins. SRP54 consists of two separable domains, a 33-kD amino-terminal domain that contains a GTP-binding site (SRP54G) and a 22-kD carboxy-terminal domain (SRP54M) containing binding sites for both the signal sequence and SRP RNA. To examine the function of the two domains in more detail, we have purified SRP54M and used it to assemble a partial SRP that lacks the amino-terminal domain of SRP54 [SRP(-54G)]. This particle recognized signal sequences in two independent assays, albeit less efficiently than intact SRP. Analysis of the signal sequence binding activity of free SRP54 and SRP54M supports the conclusion that SRP54M binds signal sequences with lower affinity than the intact protein. In contrast, when SRP(-54G) was assayed for its ability to promote the translocation of preprolactin across microsomal membranes, it was completely inactive, apparently because it was unable to interact normally with the SRP receptor. These results imply that SRP54G plays an essential role in SRP-mediated targeting of nascent chain-ribosome complexes to the ER membrane and also influences signal sequence recognition, possibly by promoting a tighter association between signal sequences and SRP54M.
Department of Biochemistry and Biophysics, School of Medicine, University of California, San Francisco 94143-0448.
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D000818:Animals; D001692:Biological Transport; D002451:Cell Compartmentation; D003432:Cross-Linking Reagents; D004721:Endoplasmic Reticulum / Q000378:metabolism; D020558:GTP Phosphohydrolases / Q000737:chemistry* / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism; D021382:Protein Sorting Signals / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000737:chemistry / Q000378:metabolism*; D018271:Signal Recognition Particle; D013329:Structure-Activity Relationship
D003432:Cross-Linking Reagents; D021382:Protein Sorting Signals; D011506:Proteins; D012261:Ribonucleoproteins; D018271:Signal Recognition Particle; D020558:GTP Phosphohydrolases