Abstract
We have investigated the formation of porphyrin intermediates by isolated barley (Hordeum vulgare) plastids incubated for 40 min with the porphyrin precursor 5-aminolevulinate and in the presence and absence of a diphenylether herbicide that blocks protoporphyrinogen oxidase, the enzyme in chlorophyll and heme synthesis that oxidizes protoporphyrinogen IX to protoporphyrin IX. In the absence of herbicide, about 50% of the protoporphyrin IX formed was found in the extraplastidic medium, which was separated from intact plastids by centrifugation at the end of the incubation period. In contrast, uroporphyrinogen, an earlier intermediate, and magnesium protoporphyrin IX, a later intermediate, were located mainly within the plastid. When the incubation was carried out in the presence of a herbicide that inhibits protoporphyrinogen oxidase, protoporphyrin IX formation by the plastids was completely abolished, but large amounts of protoporphyrinogen accumulated in the extraplastidic medium. To detect extraplastidic protoporphyrinogen, it was necessary to first oxidize it to protoporphyrin IX with the use of a herbicide-resistant protoporphyrinogen oxidase enzyme present in Escherichia coli membranes. Protoporphyrinogen is not detected by some commonly used methods for porphyrin analysis unless it is first oxidized to protoporphyrin IX. Protoporphyrin IX and protoporphyrinogen found outside the plastid did not arise from plastid lysis, because the percentage of plastid lysis, measured with a stromal marker enzyme, was far less than the percentage of these porphyrins in the extraplastidic fraction. These findings suggest that of the tetrapyrrolic intermediates synthesized by the plastids, protoporphyrinogen and protoporphyrin IX, are the most likely to be exported from the plastid to the cytoplasm. These results help explain the extraplastidic accumulation of protoporphyrin IX in plants treated with photobleaching herbicides. In addition, these findings suggest that plastids may export protoporphyrinogen or protoporphyrin IX for mitochondrial heme synthesis.
Full Text
The Full Text of this article is available as a PDF (801.9 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Bonkovsky H. L., Wood S. G., Howell S. K., Sinclair P. R., Lincoln B., Healey J. F., Sinclair J. F. High-performance liquid chromatographic separation and quantitation of tetrapyrroles from biological materials. Anal Biochem. 1986 May 15;155(1):56–64. doi: 10.1016/0003-2697(86)90224-1. [DOI] [PubMed] [Google Scholar]
- Bowyer J. R., Hallahan B. J., Camilleri P., Howard J. Mode of Action Studies on Nitrodiphenyl Ether Herbicides : II. The Role of Photosynthetic Electron Transport in Scenedesmus obliquus. Plant Physiol. 1989 Feb;89(2):674–680. doi: 10.1104/pp.89.2.674. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Camadro J. M., Matringe M., Scalla R., Labbe P. Kinetic studies on protoporphyrinogen oxidase inhibition by diphenyl ether herbicides. Biochem J. 1991 Jul 1;277(Pt 1):17–21. doi: 10.1042/bj2770017. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Castelfranco P. A., Weinstein J. D., Schwarcz S., Pardo A. D., Wezelman B. E. The Mg insertion step in chlorophyll biosynthesis. Arch Biochem Biophys. 1979 Feb;192(2):592–598. doi: 10.1016/0003-9861(79)90130-9. [DOI] [PubMed] [Google Scholar]
- Grandchamp B., Deybach J. C., Grelier M., de Verneuil H., Nordmann Y. Studies of porphyrin synthesis in fibroblasts of patients with congenital erythropoietic porphyria and one patient with homozygous coproporphyria. Biochim Biophys Acta. 1980 May 22;629(3):577–586. doi: 10.1016/0304-4165(80)90163-4. [DOI] [PubMed] [Google Scholar]
- Henderson M. J. Thin-layer chromatography of free porphyrins for diagnosis of porphyria. Clin Chem. 1989 Jun;35(6):1043–1044. [PubMed] [Google Scholar]
- Jacobs J. M., Jacobs N. J., De Maggio A. E. Protoporphyrinogen oxidation in chloroplasts and plant mitochondria, a step in heme and chlorophyll synthesis. Arch Biochem Biophys. 1982 Oct 1;218(1):233–239. doi: 10.1016/0003-9861(82)90341-1. [DOI] [PubMed] [Google Scholar]
- Matringe M., Camadro J. M., Block M. A., Joyard J., Scalla R., Labbe P., Douce R. Localization within chloroplasts of protoporphyrinogen oxidase, the target enzyme for diphenylether-like herbicides. J Biol Chem. 1992 Mar 5;267(7):4646–4651. [PubMed] [Google Scholar]
- Matringe M., Camadro J. M., Labbe P., Scalla R. Protoporphyrinogen oxidase as a molecular target for diphenyl ether herbicides. Biochem J. 1989 May 15;260(1):231–235. doi: 10.1042/bj2600231. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Matringe M., Camadro J. M., Labbe P., Scalla R. Protoporphyrinogen oxidase inhibition by three peroxidizing herbicides: oxadiazon, LS 82-556 and M&B 39279. FEBS Lett. 1989 Mar 13;245(1-2):35–38. doi: 10.1016/0014-5793(89)80186-3. [DOI] [PubMed] [Google Scholar]
- Pardo A. D., Chereskin B. M., Castelfranco P. A., Franceschi V. R., Wezelman B. E. ATP requirement for mg chelatase in developing chloroplasts. Plant Physiol. 1980 May;65(5):956–960. doi: 10.1104/pp.65.5.956. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Porra R. J., Falk J. E. The enzymic conversion of coproporphyrinogen 3 into protoporphyrin 9. Biochem J. 1964 Jan;90(1):69–75. doi: 10.1042/bj0900069. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Porra R. J., Lascelles J. Studies on ferrochelatase. The enzymic formation of haem in proplastids, chloroplasts and plant mitochondria. Biochem J. 1968 Jun;108(2):343–348. doi: 10.1042/bj1080343. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Smith A. G. Subcellular localization of two porphyrin-synthesis enzymes in Pisum sativum (pea) and Arum (cuckoo-pint) species. Biochem J. 1988 Jan 15;249(2):423–428. doi: 10.1042/bj2490423. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Witkowski D. A., Halling B. P. Inhibition of plant protoporphyrinogen oxidase by the herbicide acifluorfen-methyl. Plant Physiol. 1989 Aug;90(4):1239–1242. doi: 10.1104/pp.90.4.1239. [DOI] [PMC free article] [PubMed] [Google Scholar]