Ribosome-independent regulation of translocon composition and Sec61alpha conformation.

Journal Article (Journal Article)

In this study, the contributions of membrane-bound ribosomes to the regulation of endoplasmic reticulum translocon composition and Sec61alpha conformation were examined. Following solubilization of rough microsomes (RM) with digitonin, ribosomes co-sedimented in complexes containing the translocon proteins Sec61alpha, ribophorin I, and TRAPalpha, and endoplasmic reticulum phospholipids. Complexes of similar composition were identified in digitonin extracts of ribosome-free membranes, indicating that the ribosome does not define the composition of the digitonin-soluble translocon. Whereas in digitonin solution a highly electrostatic ribosome-translocon junction is observed, no stable interactions between ribosomes and Sec61alpha, ribophorin I, or TRAPalpha were observed following solubilization of RM with lipid-derived detergents at physiological salt concentrations. Sec61alpha was found to exist in at least two conformational states, as defined by mild proteolysis. A protease-resistant form was observed in RM and detergent-solubilized RM. Removal of peripheral proteins and ribosomes markedly enhanced the sensitivity of Sec61alpha to proteolysis, yet the readdition of inactive ribosomes to salt-washed membranes yielded only modest reductions in protease sensitivity. Addition of sublytic concentrations of detergents to salt-washed RM markedly decreased the protease sensitivity of Sec61alpha, indicating that a protease-resistant conformation of Sec61alpha can be conferred in a ribosome-independent manner.

Full Text

Duke Authors

Cited Authors

  • Potter, MD; Nicchitta, CV

Published Date

  • January 21, 2000

Published In

Volume / Issue

  • 275 / 3

Start / End Page

  • 2037 - 2045

PubMed ID

  • 10636907

International Standard Serial Number (ISSN)

  • 0021-9258

Digital Object Identifier (DOI)

  • 10.1074/jbc.275.3.2037


  • eng

Conference Location

  • United States