Mechanisms of integration of de novo-synthesized polypeptides into membranes: signal-recognition particle is required for integration into microsomal membranes of calcium ATPase and of lens MP26 but not of cytochrome b5.

Abstract

The in vitro integration into dog pancreas microsomal membranes of 3 integral membrane proteins that were synthesized do novo in a wheat germ cell-free translation system was investigated: Ca-ATPase of rabbit sarcomplasmic reticulum, MP26 of bovine lens fiber plasma membrane and rat liver cytochrome b5. Biosynthetically these proteins show a common feature in that they are synthesized without a transient NH2-terminal signal sequence. Two of these proteins, ATPase and MP26, were shown to require the recently discovered signal-recognition particle (SRP) for integration. By this criterion, therefore, they each contain at least 1 uncleaved signal sequence. However, the uncleaved signal sequence(s) of these 2 proteins did not induce the characteristic SRP-mediated translation arrest that was previously shown for a cleaved signal sequence. Unlike ATP-ase and MP26, cytochrome b5 did not require SRP for integration into microsomal membrane. Thus, the distinction between an insertion sequence (specifying unassisted and opportunistic integration into any exposed membrane) and a signal sequence (directing integration into a specific membrane by a receptor-mediated mechanism) is a valid one. By assaying for SRP dependence, the 2 mechanisms of integration can now be experimentally distinguished.