Protochlorophyll Biosynthesis in a Cell-free System from Higher Plants

A cell free system prepared from etiolated cucumber (Cucumis sativus, L) in tris-sucrose buffer is able to incorporate delta-aminolevulinic acid-4- (14)C into the two components of protochlorophyll: protochlorophyllide and protochlorophyllide ester. The activity is associated with the etioplasts. Optimal incorporation is obtained at pH 7.7. For the formation of protochlorphyllide ester, oxygen, reduced glutathione, methyl alcohol, magnesium, inorganic phosphate, and nicotinamide adenine dinucleotide are required. For the formation of (14)C-protochlorophyllide, adenosine triphosphate, and coenzyme A are required in addition to the above. The requirement for methyl alcohol is highly specific, and the methyl group appears to be incorporated into the protochlorophyll molecules. A biosynthetic scheme resulting in the parallel production of (14)C-protochlorophyllide and (14)C-protochlorophyllide ester from (14)C-Mg protoporphyrin monoester is presented.