Hamdan F F, Ribeiro P
Institute of Parasitology, McGill University, Quebec H9X 3V9, Canada.
J Biol Chem. 1999 Jul 30;274(31):21746-54. doi: 10.1074/jbc.274.31.21746.
A cDNA (Schistosoma mansoni tryptophan hydroxylase; SmTPH) encoding a protein homologous to tryptophan hydroxylase, the enzyme that catalyzes the rate-limiting step in the biosynthesis of serotonin, was cloned from the human parasite Schistosoma mansoni. Bacterial expression of SmTPH as a histidine fusion protein produced soluble active enzyme, which was purified to apparent homogeneity and a final specific activity of 0.17 micromol/min/mg of protein. The purified enzyme was found to be a tetramer of approximately 240 kDa with a subunit size of 58 kDa. Several of the biochemical and kinetic properties of SmTPH were similar to those of mammalian tryptophan hydroxylase. Unlike the mammalian enzyme, however, SmTPH was found to be stable at 37 degrees C, its t((1)/(2)) being nearly 23 times higher than that of a similarly expressed rabbit tryptophan hydroxylase. A semiquantitative reverse transcription polymerase chain reaction showed that the level of SmTPH mRNA in a larval stage of the parasite (cercaria) is 2.5 times higher than in adult S. mansoni, suggesting possible differences in the level of enzyme expression between the two developmental stages. This study demonstrates for the first time the presence of a functional tryptophan hydroxylase in a parasitic helminth and further suggests that the parasites are capable of synthesizing serotonin endogenously.
从人体寄生虫曼氏血吸虫中克隆出一种互补DNA（曼氏血吸虫色氨酸羟化酶；SmTPH），其编码的蛋白质与色氨酸羟化酶同源，色氨酸羟化酶是催化血清素生物合成限速步骤的酶。SmTPH作为组氨酸融合蛋白在细菌中表达产生可溶性活性酶，该酶被纯化至表观均一，最终比活性为0.17微摩尔/分钟/毫克蛋白质。发现纯化后的酶是一种约240 kDa的四聚体，亚基大小为58 kDa。SmTPH的一些生化和动力学特性与哺乳动物色氨酸羟化酶相似。然而，与哺乳动物酶不同的是，SmTPH在37℃下稳定，其半衰期比同样表达的兔色氨酸羟化酶高出近23倍。半定量逆转录聚合酶链反应表明，寄生虫幼虫期（尾蚴）的SmTPH mRNA水平比成年曼氏血吸虫高2.5倍，这表明两个发育阶段的酶表达水平可能存在差异。这项研究首次证明了寄生蠕虫中存在功能性色氨酸羟化酶，并进一步表明寄生虫能够内源性合成血清素。
