Translocation of secretory proteins across the microsomal membrane occurs through an environment accessible to aqueous perturbants.
We have characterized the association of a nascent secretory protein with the microsomal membrane at two distinct stages in cell-free synthesis and translocation. Stage one corresponded to a nascent chain of approximately 70 residues generated via elongation arrest by the signal recognition particle (SRP). Binding to microsomal membranes occurred independently of chain elongation and required SRP receptor. Following binding, the 70-mer remained attached to the membrane after extraction of the ribosome. However, protein denaturants (4 M urea or alkaline pH) extracted the 70-mer from the membrane. Stage two of synthesis corresponded to nascent chains of approximately 158 residues generated by oligonucleotide-mediated hybrid arrest of translation. Again, these partially translocated nascent chains were extracted by 4 M urea. Therefore, the initial interaction of the signal sequence with the membrane as well as subsequent chain conductance occur in a microenvironment that is accessible to aqueous reagents. Thus, both processes probably require integral membrane proteins.
Gilmore
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Blobel
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D000818:Animals; D002474:Cell-Free System; D004285:Dogs; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins / Q000502:physiology*; D008861:Microsomes / Q000378:metabolism*; D010441:Peptide Chain Elongation, Translational; D010443:Peptide Chain Termination, Translational; D010446:Peptide Fragments / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism* / Q000494:pharmacology; D011132:Polyribosomes / Q000378:metabolism; D011388:Prolactin / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism*; D021382:Protein Sorting Signals; D011691:Puromycin / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear; D018000:Receptors, Peptide; D014508:Urea / Q000494:pharmacology
D008565:Membrane Proteins; D010446:Peptide Fragments; D010455:Peptides; D011498:Protein Precursors; D021382:Protein Sorting Signals; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; D018000:Receptors, Peptide; C036318:signal peptide receptor; D011691:Puromycin; C017671:preprolactin; D014508:Urea; D011388:Prolactin