Chlorophyll (Chl) b is essential for maximal capacity of photosynthesis in green algae and plants. Along with broadening the absorbance spectrum, Chl b controls the translocation of light-harvesting complex apoproteins across the chloroplast envelope inner membrane and stabilizes light-harvesting complexes in thylakoid membranes. In the absence of Chl b, these Chl-binding proteins are retracted into the cytosol and degraded. A protein designated chlorophyllide (Chlide) a oxygenase (CAO) is required for Chl b synthesis. Conversion in vitro of Chlide a to Chlide b by purified recombinant CAO occurs at a low rate. In contrast, protochlorophyllide (Pchlide) a is converted rapidly and quantitatively to Chlide b by CAO in a membrane fraction from cells of Chlamydomonas reinhardtii y-1 when incubated in the presence of polyaromatic m-phenanthroline. This is surprising because Pchlide a per se is not a substrate for CAO. A retrospective analysis of our previous data suggests that phenanthroline mimics Chlide a and pairs with Pchlide a to lower its redox potential, thereby activating Pchlide a as a substrate. A conserved tyrosine residue occurs as a stable radical in CAO and initiates the transformation of the 7-methyl group of Pchlide a to the 7-formyl group of Pchlide b. Replacing this tyrosine with alanine abolished CAO activity. We conclude that Pchlide b is synthesized from Pchlide a by an unusual, radical-mediated/based mechanism that directly involves molecular oxygen. In vivo, subsequent reduction of the C17-C18 double bond by a light-independent Pchlide reductase activity would be facilitated by the electronegative carbonyl of the 7-formyl group to generate Chlide b.
Hoober et al. (Thu,) studied this question.