Phototransformation of protochlorophyll(ide) by a proplastid fraction from Euglena

The phototransformation of protochlorophyll(ide) (Pchl(ide)) to chlorophyll(ide) (Chl(ide)) can be demonstrated in a proplastid fraction from Euglena gracilis Klebs var. bacillaris Cori if appropriate conditions are employed. Pigments were measured fluorometrically in acetone extracts of cell or organelles. Pchl(ide) and the phototransformation to Chl(ide) are at their highest levels in cells grown in darkness on normal or low vitamin B₁₂-containing medium (pH 3.5) to the late exponential phase (1.2–1.4 × 10⁶ cells ml^?1). Late exponential cells on low B¹² medium yield a proplastid fraction that contains Pchl(ide) which is phototransformed to Chl(ide) when illuminated with red light (5.6 W m^?2 for 4 min) in the presence of 10 mM Hepes, 20 mM TES, 0.5 mM potassium phosphate (pH 7.4), 70 mM sorbitol, 5 mM DTT, 5 mM ATP, 5 mM fructose-1, 6-bisphosphate, 10 mM malate and 2 mM MgCl²; intact organelles appear to be involved since deletion of osmoticum gives a lower activity, and addition of NAD(P)H is without effect. Phototransformation of Pchl(ide) to Chl(ide) in red light shows Bunsen-Roscoe reciprocity between fluence rate and duration of illumination. Although mitochondria are present, they do not appear to be involved since inhibitors of respiration and uncouplers of oxidative phosphorylation fail to block the phototransformation. The percentage phototransformation of Pchl(ide) to Chl(ide) in late exponential normal B¹² cells is 61 ± 10, and is 52 ± 3 in low B¹² cells. About 67% of the activity in low B¹² cells is recovered in the proplastid fraction incubated with the complete incubation mixture in saturating light. In both types of cells and in the proplastid fraction, the stoichiometry of conversion of Pchl(ide) to Chl(ide) is about 1:1 (mol/mol).