The distribution of radioactive peptides synthesized by polysomes and ribosomal subunits combined in vitro with microsomal membranes

Abstract

1. Rat liver endoplasmic reticulum membranes were conditioned in various ways to bind exogenous polysomes in vitro. The puromycin-mediated release of nascent peptides from polysomes bound to such membranes was then studied, in efforts to determine whether the complexes functioned as secretory units, directing released peptides toward the inside of the membrane vesicle rather than outward toward the incubation medium. The distribution of released peptides between the incubation medium and a detergent-soluble fraction, which represents the vesicle membrane and contents, was examined. 2. The following combinations of ribosomes and microsomal membranes were tested: (a) The hormone-mediated binding of free polysomes to smooth endoplasmic reticulum membranes; (b) the binding of polysomes to membranes conditioned with citrate and pyrophosphate, with EDTA and ribonuclease, or with puromycin at high concentrations of KCl; and (c) the interaction of ribosomal subunits and polyuridylic acid with puromycin-KCl-conditioned membranes. 3. There is no evidence of vectorial transport of nascent peptides across membranes in in vitro recombination systems. It is concluded that these systems do not duplicate the type of binding observed in naturally occurring rough-surfaced endoplasmic reticulum.