Protein targeting across the three membranes of the Euglena chloroplast envelope

Abstract

A system has been developed for the import in vitro of precursor proteins into Euglena chloroplasts, which have three envelope membranes. Preparation of functional chloroplasts with intact envelope membranes has been optimized. Import of the precursor (50 kDa) for the tetrapyrrole biosynthesis enzyme porphobilinogen deaminase (PBGD), and processing to the mature size (40 kDa), occurred at 25 °C in the light and the presence of ATP, with an estimated efficiency of 62%. Pretreatment of the chloroplasts with proteases abolished this import, suggesting the involvement of specific protein receptors. The presequence of PBGD was found to be cleaved by Escherichia coli leader peptidase to an interme..