Nakajima K, Hashimoto T, Yamada Y
Department of Agricultural Chemistry, Faculty of Agriculture, Kyoto University, Japan.
Proc Natl Acad Sci U S A. 1993 Oct 15;90(20):9591-5. doi: 10.1073/pnas.90.20.9591.
In the biosynthetic pathway of tropane alkaloids, tropinone reductase (EC 1.1.1.236) (TR)-I and TR-II, respectively, reduce a common substrate, tropinone, stereospecifically to the stereoisomeric alkamines tropine and pseudotropine (psi-tropine). cDNA clones coding for TR-I and TR-II, as well as a structurally related cDNA clone with an unknown function, were isolated from the solanaceous plant Datura stramonium. The cDNA clones for TR-I and TR-II encode polypeptides containing 273 and 260 amino acids, respectively, and when these clones were expressed in Escherichia coli, the recombinant TRs showed the same strict stereospecificity as that observed for the native TRs that had been isolated from plants. The deduced amino acid sequences of the two clones showed an overall identity of 64% in 260-amino acid residues and also shared significant similarities with enzymes in the short-chain, nonmetal dehydrogenase family. Genomic DNA-blot analysis detected the TR-encoding genes in three tropane alkaloid-producing solanaceous species but did not detect them in tobacco. We discuss how the two TRs may have evolved to catalyze the opposite stereospecific reductions.
在托烷生物碱的生物合成途径中，托品酮还原酶（EC 1.1.1.236）（TR）-I和TR-II分别将共同底物托品酮立体定向还原为立体异构的烷胺托品和假托品（ψ-托品）。从茄科植物曼陀罗中分离出编码TR-I和TR-II的cDNA克隆，以及一个功能未知但结构相关的cDNA克隆。TR-I和TR-II的cDNA克隆分别编码含有273和260个氨基酸的多肽，当这些克隆在大肠杆菌中表达时，重组TRs表现出与从植物中分离出的天然TRs相同的严格立体特异性。这两个克隆推导的氨基酸序列在260个氨基酸残基中总体一致性为64%，并且与短链、非金属脱氢酶家族中的酶也有显著相似性。基因组DNA印迹分析在三种产生托烷生物碱的茄科植物中检测到了TR编码基因，但在烟草中未检测到。我们讨论了这两种TRs可能是如何进化以催化相反的立体特异性还原反应的。
