Protein synthesis in isolated spinach chloroplasts: Comparison of light-driven and ATP-driven synthesis

Abstract

A comparison has been made of the optimal concentrations of Mg 2+ and K + ions necessary for both light-driven protein synthesis in intact spinach chloroplasts and for ATP-driven protein synthesis in broken chloroplasts, and the products of the two systems have been compared by polyacrylamide gel electrophoresis. Light-driven incorporation of amino acids into polypeptides in intact chloroplasts assayed in buffer systems containing sucrose or sorbitol as the osmoticum is inhibited by the addition of Mg 2+, the effect being most marked at low concentrations (less than 40 m m) of KCl. On the other hand, chloroplasts suspended in 0.2 m KCl as osmoticum require Mg 2+ (3 m m) for optimal light-driven protein-synthesizing activity. Incorporation of amino acids by broken chloroplasts in the dark, supplemented with ATP and GTP, requires 9 m m Mg 2+ for maximum activity. A requirement for monovalent cations is best filled by K + (approx 30 m m) in the case of the light-driven, intact chloroplast system whereas, in the ATP-driven, broken chloroplast system, NH 4 + (approx 80 m m) gave the highest activity. Autoradiographs of Na dodecyl sulfate-polyacrylamide gels of the products from both the light-driven, intact chloroplasts and from the ATP-driven, broken chloroplasts reveal qualitatively similar patterns. There are at least four radioactive polypeptides in the soluble protein fraction the dominant product being coincident with the large subunit of Fraction 1 protein. In the membrane fraction at least nine discrete products can be resolved.