Witty M, Jones R M, Robb M S, Jordan P M, Smith A G
Department of Plant Sciences, University of Cambridge, UK.
Planta. 1996;199(4):557-64. doi: 10.1007/BF00195187.
A recombinant plasmid, pArab8, harbouring the cDNA encoding the mature form of the tetrapyrrole synthesis enzyme porphobilinogen deaminase (EC 4.3.1.8; also known as hydroxymethylbilane synthase) from Arabidopsis thaliana (L.) Heynh. has been constructed, and used to transform Escherichia coli. The porphobilinogen deaminase protein from Arabidopsis was overexpressed in this strain, and purified to homogeneity (3000-fold) with a yield of 20%. Antibodies were raised against the purified plant enzyme, and used in Western blot analysis, immunoprecipitation of enzyme activity and immuno-gold electron microscopy. The results indicate that the enzyme is confined to plastids in both leaves and roots. The implications of this finding for plant tetrapyrrole synthesis are discussed.
已构建了一种重组质粒pArab8，其含有编码来自拟南芥（L.）Heynh.的四吡咯合成酶胆色素原脱氨酶（EC 4.3.1.8；也称为羟甲基胆色素合酶）成熟形式的cDNA，并用于转化大肠杆菌。来自拟南芥的胆色素原脱氨酶蛋白在该菌株中过表达，并以20%的产率纯化至均一性（3000倍）。制备了针对纯化的植物酶的抗体，并用于蛋白质免疫印迹分析、酶活性的免疫沉淀和免疫金电子显微镜检查。结果表明，该酶在叶片和根中均局限于质体中。讨论了这一发现对植物四吡咯合成的意义。
