ENVELOPE MEMBRANE-PROTEINS THAT INTERACT WITH CHLOROPLASTIC PRECURSORPROTEINS

The post-translational transport of cytoplasmically synthesized precur sor proteins into chloroplasts requires proteins in the envelope membr anes. To identify some of these proteins, label transfer cross-linking was performed using precursor to the small subunit of ribulose-1,5-bi sphosphate carboxylase (prSSU) that was blocked at an early stage of t he transport process. Two envelope proteins were identified: an 86-kD protein and a 75-kD protein, both present in the outer membrane. Label ing of both proteins required prSSU and could not be accomplished with SSU lacking a transit peptide. Labeling of the 75-kD protein occurred only when low levels of ATP were present, whereas labeling of the 86- kD protein occurred in the absence of exogenous ATP. Although both lab eled proteins were identified as proteins of the outer envelope membra ne, the labeled form of the 75-kD protein could only be detected in fr actions containing mixed envelope membranes. Based on these observatio ns, we propose that prSSU first binds in an ATP-independent fashion to the 86-kD protein. The energy-requiring step is association with the 75-kD protein and assembly of a translocation contact site between the inner and outer membrane of the chloroplastic envelope.