Secretion requires a cytoplasmically disposed sulphydryl of the RER membrane

Abstract

The capacity of the rough endoplasmic reticulum (RER) membrane of eukaryotic cells to translocate nascent presecretory proteins from the cytosol to the intracisternal space is preserved on cell fractionation and can be assayed in vitro¹. Two attempts to characterize this translocation activity have been reported. Warren and Dobberstein² reported that microsomal membranes can be depleted of their translocation activity by extraction with a solution of high ionic strength (500 mM KCl) and that activity can be restored to the depleted membranes by re-addition of the salt extract. On the other hand, Walter et al.³ reported that KCl extraction of the microsomal membrane does not result in complete depletion of its translocation activity. However, mild trypsinization of the microsomal membrane released a tryptic fragment(s) from the membrane which, when recombined with a tryptically inactivated membrane fraction, restored translocation activity³. We now show that both the trypsin and the KCl extracted factors, but not the membrane-integrated remainder of the translocation apparatus, contain at least one sulphydryl group that is essential for activity.