Localization within chloroplasts of protoporphyrinogen oxidase, the target enzyme for diphenylether-like herbicides.
Plant protoporphyrinogen oxidase is of particular interest since it is the last enzyme of the common branch for chlorophyll and heme biosynthetic pathways. In addition, it is the target enzyme for diphenyl ether-type herbicides, such as acifluorfen. Two distinct methods were used to investigate the localization of this enzyme within Percoll-purified spinach chloroplasts. We first assayed the enzymatic activity by spectrofluorimetry and we analyzed the specific binding of the herbicide acifluorfen, using highly purified chloroplast fractions. The results obtained give clear evidence that chloroplast protoporphyrinogen oxidase activity is membrane-bound and is associated with both chloroplast membranes, i.e. envelope and thylakoids. Protoporphyrinogen oxidase specific activity was 7-8 times higher in envelope membranes than in thylakoids, in good agreement with the number of [3H]acifluorfen binding sites in each membrane system: 21 and 3 pmol/mg protein, respectively, in envelope membranes and thylakoids. On a total activity basis, 25% of protoporphyrinogen oxidase activity were associated with envelope membranes. The presence of protoporphyrinogen oxidase in chloroplast envelope membranes provides further evidence for a role of this membrane system in chlorophyll biosynthesis. In contrast, the physiological significance of the enzyme associated with thylakoids is still unknown, but it is possible that thylakoid protoporphyrinogen oxidase could be involved in heme biosynthesis.
Matringe
M
M
URA Centre National de la Recherche Scientifique (CNRS) No. 576, Département de Biologie Moléculaire et Structurale, Grenoble, France.
Camadro
J M
JM
Block
M A
MA
Joyard
J
J
Scalla
R
R
Labbe
P
P
Douce
R
R
D002736:Chloroplasts / Q000201:enzymology*; D006418:Heme / Q000096:biosynthesis; D006540:Herbicides / Q000378:metabolism*; D007425:Intracellular Membranes / Q000201:enzymology; D009578:Nitrobenzenes / Q000378:metabolism*; D009579:Nitrobenzoates / Q000378:metabolism*; D010088:Oxidoreductases / Q000378:metabolism*; D044925:Oxidoreductases Acting on CH-CH Group Donors; D050768:Protoporphyrinogen Oxidase
D006540:Herbicides; D009578:Nitrobenzenes; D009579:Nitrobenzoates; D006418:Heme; C064882:acifluorfen-methyl; D010088:Oxidoreductases; D044925:Oxidoreductases Acting on CH-CH Group Donors; D050768:Protoporphyrinogen Oxidase; C018425:acifluorfen