An ATP-binding membrane protein is required for protein translocation across the endoplasmic reticulum membrane.
The role of nucleotides in providing energy for polypeptide transfer across the endoplasmic reticulum (ER) membrane is still unknown. To address this question, we treated ER-derived mammalian microsomal vesicles with a photoactivatable analogue of ATP, 8-N3ATP. This treatment resulted in a progressive inhibition of translocation activity. Approximately 20 microsomal membrane proteins were labeled by [alpha 32P]8-N3ATP. Two of these were identified as proteins with putative roles in translocation, alpha signal sequence receptor (SSR), the 35-kDa subunit of the signal sequence receptor complex, and ER-p180, a putative ribosome receptor. We found that there was a positive correlation between inactivation of translocation activity and photolabeling of alpha SSR. In contrast, our data demonstrate that the ATP-binding domain of ER-p180 is dispensable for translocation activity and does not contribute to the observed 8-N3ATP sensitivity of the microsomal vesicles.
Zimmerman
D L
DL
Department of Biochemistry and Biophysics University of California, Medical School San Francisco 94143-0448.
Walter
P
P
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism*; D000345:Affinity Labels; D000818:Animals; D001386:Azides; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D008861:Microsomes / Q000378:metabolism; D021382:Protein Sorting Signals / Q000378:metabolism
D000345:Affinity Labels; D001386:Azides; D002352:Carrier Proteins; D008565:Membrane Proteins; D021382:Protein Sorting Signals; C017040:8-azidoadenosine 5'-triphosphate; D000255:Adenosine Triphosphate