Interaction between membrane properties and protein synthesis: In vitro synthesis of membrane proteins during erythropoiesis

Abstract

Summary Membrane protein synthesis was investigated by incubating rabbit reticulocytes, in vitro, with radioactive amino acids. The kinetics of membrane protein synthesis showed linear incorporation for approx. 15 min, after which there was only a slight increase in incorporation. On the other hand, intracellular protein synthesis was linear for an incubation period of 60 min. Membranes isolated from such rabbit reticulocytes were analysed on sodium dodecyl sulfate (SDS)-polyacrylamide gels. Two major radioactive bands were found in the 50–60 000 D region, whilst another labelled band had a molecular weight of 43 000 D. This latter band had an electrophoretic mobility identical with rabbit muscle actin (and chick brain actin), when run on one-dimensional SDS polyacrylamide gels. Absolute identity between rabbit brain actin and a newly synthesized reticulocyte membrane protein was shown by comigration on a two-dimensional (first dimension isoelectric focusing and second dimension SDS gel) electrophoresis system. Another band that was radioactively labelled was found to have a molecular weight of approx. 32 000 D. Separation of reticulocytes into different age groups showed that young reticulocytes synthesized a membrane protein species that was not radioactively labelled in the old reticulocyte population.