UMMS Affiliation

Department of Biochemistry and Molecular Biology

Date

7-1-1991

Document Type

Article

Subjects

*Calcium-Binding Proteins; Cell-Free System; Cross-Linking Reagents; Endoplasmic Reticulum; Ethylmaleimide; *Membrane Glycoproteins; Membrane Proteins; Peptides; RNA, Transfer; RNA, Transfer, Amino Acyl; Receptors, Cell Surface; *Receptors, Cytoplasmic and Nuclear; *Receptors, Peptide; Ribonucleoproteins; Signal Recognition Particle; Succinimides; Viral Envelope Proteins

Disciplines

Biochemistry | Cell Biology | Molecular Biology

Abstract

We have used the homobifunctional cross-linking reagent disuccinimidyl suberate (DSS) to identify proteins that are adjacent to nascent polypeptides undergoing translocations across mammalian rough ER. Translocation intermediates were assembled by supplementing cell free translations of truncated mRNAs with the signal recognition particle (SRP) and microsomal membrane vesicles. Two prominent cross-linked products of 45 and 64 kD were detected. The 64-kD product was obtained when the cell free translation contained SRP, while formation of the 45-kD product required both SRP and translocation competent microsomal membrane vesicles. In agreement with previous investigators, we suggest that the 64-kD product arises by cross-linking of the nascent polypeptide to the 54-kD subunit of SRP. The 45-kD product resists alkaline extraction from the membrane, so we conclude that the 11-kD nascent polypeptide has been crosslinked to an integral membrane protein of approximately 34 kD (imp34). The cross-linked product does not bind to ConA Sepharose, nor is it sensitive to endoglycosidase H digestion; hence imp34 is not identical to the alpha or beta subunits of the signal sequence receptor (SSR). We propose that imp34 functions in concert with SSR to form a translocation site through which nascent polypeptides pass in traversing the membrane bilayer of the rough endoplasmic reticulum.

Rights and Permissions

Citation: J Cell Biol. 1991 Jul;114(1):21-33.

Related Resources

Link to Article in PubMed

Journal Title

The Journal of cell biology

PubMed ID

1646822

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