Abstract
It is widely accepted that plastids have evolved from free-living cyanobacteria. In the process of endosymbiosis, most of genes were relocated from ancestral plastids to the host nucleus. Most nuclear-encoded plastid proteins are cytosolically synthesized with N-terminal transit peptide (TP). It has been experimentally demonstrated that several TPs lack defined secondly structure. Such unstructured proteins are also known to be unstable in bacterial cells.
Here, we hypothesized that stabilization of unstructured proteins in eukaryotic cytoplasm would be prerequisite for acquisition of TP. To assess this hypothesis, we examined the fate of a TP of RbcS fused to GFP in E.coli. Addition of N-terminal stable tag allowed us to purify the fusion protein. After removal of the N-terminal tag by specific protease, the TP-GFP was rapidly degraded in the presence of E.coli cell extract. This degradation was inhibited by addition of o-phenanthroline, suggesting that metalloproteases mainly degrade the TP. Based on these results, it is assumed that such protease activity would be lost in eukaryotic cytoplasm before the acquisition of TP.
