Tryptic digestion of membrane proteins synthesized on chloroplast ribosomes in Chlamydomonas reinhardtii.

Chloroplast membranes of the green alga Chlamydomonas reinhardtii have been isolated from cells incubated in the presence of cycloheximide. The proteins produced under these conditions are formed on chloroplastic ribosomes. Fluorography of the polypeptides separated on polyacrylamide gels shows ten major protein species associated with the membranes that are produced on chloroplast ribosomes. These membranes were subjected to differential proteolysis. The change in electrophoretic mobility of the membrane proteins after controlled digestion indicates that proteins of molecular weight 55 000, 43 000 and 30 000 are either buried in the membrane or protected by association with other proteins. Most, if not all, of the chloroplast-synthesized thylakoid proteins contain trypsin-sensitive sites accessible from the stromal (out)-side of the isolated membrane vesicle. Thus, most chloroplast-synthesized membrane proteins are asymmetrically distributed in the thylakoid.