The overall objective of the proposed research is to elucidate the mechanism of protein biosynthesis in cell organelles such as mitochondria and chloroplasts and to compare this system to prokaryotic and eukaryotic cytoplasmic protein synthesizing systems. We will seek to identify and characterize the organelle factors required for the translation of natural messenger RNA using factors from the chloroplasts of Euglena gracilis as a model system. Our initial goal will be to establish an in vitro system from chloroplasts which will translate both synthetic and natural messenger RNAs. We then propose to isolate, purify and characterize the organelle elongation factors and to compare them to prokaryotic and cytoplasmic elongation factors. Our second objective will be to purify the initiation factors and to define their roles in the initiation process. This will permit us to compare the process of chain initiation in cell organelles, bacteria and the cytoplasm of eukaryotic cells. A long-term objective is to develop a defined in vitro system in which to study the regulation of organelle protein biosynthesis and its integration into the complex metabolism of the eukaryotic cell.