Camadro J M, Matringe M, Thome F, Brouillet N, Mornet R, Labbe P
Laboratoire de Biochimie des Porphyrines, Institut Jacques Monod, CNRS, Université Paris 7, France.
Eur J Biochem. 1995 May 1;229(3):669-74. doi: 10.1111/j.1432-1033.1995.tb20512.x.
Diphenylether-type herbicides are extremely potent inhibitors of protoporphyrinogen oxidase, a membrane-bound enzyme involved in the heme and chlorophyll biosynthesis pathways. Tritiated acifluorfen and a diazoketone derivative of tritiated acifluorfen were specifically bound to a single class of high-affinity binding sites on yeast mitochondrial membranes with apparent dissociation constants of 7 nM and 12.5 nM, respectively. The maximum density of specific binding sites, determined by Scatchard analysis, was 3 pmol.mg-1 protein. Protoporphyrinogen oxidase specific activity was estimated to be 2500 nmol protoporphyrinogen oxidized h-1.mol-1 enzyme. The diazoketone derivative of tritiated acifluorfen was used to specifically photolabel yeast protoporphyrinogen oxidase. The specifically labeled polypeptide in wild-type mitochondrial membranes had an apparent molecular mass of 55 kDa, identical to the molecular mass of the purified enzyme. This photolabeled polypeptide was not detected in a protoporphyrinogen-oxidase-deficient yeast strain, but the membranes contained an equivalent amount of inactive immunoreactive protoporphyrinogen oxidase protein.
二苯醚类除草剂是原卟啉原氧化酶的极强抑制剂，原卟啉原氧化酶是一种参与血红素和叶绿素生物合成途径的膜结合酶。氚标记的三氟羧草醚及其氚标记的重氮酮衍生物特异性结合于酵母线粒体膜上的一类单一高亲和力结合位点，其表观解离常数分别为7 nM和12.5 nM。通过Scatchard分析确定的特异性结合位点的最大密度为3 pmol·mg-1蛋白质。原卟啉原氧化酶的比活性估计为2500 nmol原卟啉原氧化·h-1·mol-1酶。氚标记的三氟羧草醚的重氮酮衍生物用于特异性光标记酵母原卟啉原氧化酶。野生型线粒体膜中特异性标记的多肽的表观分子量为55 kDa，与纯化酶的分子量相同。在原卟啉原氧化酶缺陷型酵母菌株中未检测到这种光标记的多肽，但膜中含有等量的无活性免疫反应性原卟啉原氧化酶蛋白。
