Processing of a chloroplast-translated membrane protein in vivo. Analysis of the rapidly synthesized 32 000-dalton shield protein and its precursor in Spirodela oligorrhiza.

The 32 000-dalton (Da) shield protein regulating electron transport in Spirodela oligorrhiza is an integral chloroplast membrane polypeptide. It is rapidly synthesized, constituting a major chloroplast-translation product in vivo. Following in vitro translation of spirodela chloroplast RNA in a wheat germ system, a 33 500-Da polypeptide is produced. Synthesis of a 33 500-Da protein, associated with the chloroplast membrane, is also seen in vivo, within 2 min of pulse-labeling spirodela with radioactive amino acids. Comparative analyses among these polypeptides reveal: (a) all three are deficient in lysine residues; (b) the two 33 500-Da species have indistinguishable partial proteolytic digestion patterns while that for the 32 000-Da protein differs only slightly from them; (c) radioactivity from the 33 500-Da polypeptide is rapidly chased in vivo into the 32 000-Da protein, even in the presence of protein synthesis inhibitors. These results show the 33 500-Da proteins synthesized in vitro and in vivo to be the precursor form of the 32 000-Da shield protein in spirodela, with processing commencing only after completion of the precursor polypeptide chain and insertion into the membrane.