Truncations of a secretory protein define minimum lengths required for binding to signal recognition particle and translocation across the endoplasmic reticulum membrane.

Nascent preproinsulin interacts with endoplasmic reticulum membranes after approximately 70-80 residues of the 116-amino acid precursor are polymerized (Eskridge, E. M., and Shields, D. (1983) J. Biol. Chem. 258, 11487-11491). To understand the relationship between the size of a nascent presecretory polypeptide and the efficiency of its translocation across the endoplasmic reticulum membrane, recombinant DNA molecules were generated that encoded a series of preproinsulin derivatives with the same NH2 terminus as preproinsulin and progressively shorter COOH termini. The DNA was transcribed, the in vitro transcription products were translated in the wheat germ cell-free translation system, and the interaction of the resulting truncated polypeptides with signal recognition particle (SRP) and with microsomal membranes was analyzed. Truncations composed of 78 and 64 amino acids were translocated across the endoplasmic reticulum membrane, and translocation was found to be strictly co-translational and SRP-dependent. Translocation efficiency at low membrane concentrations was reduced for these truncated molecules relative to full-length preproinsulin. Most significantly, translation of the 64-residue polypeptide was arrested by SRP after only 50 amino acids were polymerized. This suggests that the initial interaction of nascent secretory proteins with SRP occurs when only 10 residues of the signal peptide protrude from the large ribosomal subunit.