Methionine aminopeptidases (MAPs), which remove the N-terminal methionine from newly synthesised proteins, are present in all life forms. Three type I MAPs and one type II MAP are encoded in the genome of Toxoplasma gondii. In this study, we found that the inducible knockdown of each type I TgMAP (TgMAP1a–c) reduced the growth and proliferation of the parasite significantly. Among them, TgMAP1c was found to be localised to the apicoplast of the parasite. Inducible knockdown of TgMAP1c led to a defect in the abundance of apicoplast-encoded transcripts, and a later reduction in the apicoplast genome copy number and loss of the apicoplast structure. This finding indicates that transcription of the apicoplast genome was impaired upon knockdown of TgMAP1c. We also found that the function of TgMAP1c in apicoplast biogenesis depends on its enzymatic domain. Expression of a recombinant protein in which the active domain of TgMAP1c was replaced with that of TgMAP1a or TgMAP1b could not restore the defective growth and replication phenotype caused by knockdown of TgMAP1c, indicating that these three enzymes have distinct substrate preferences. An in vitro analysis also revealed that TgMAP1c is an active enzyme that acts specifically on the substrate H-Met-p-NA. In addition, inducible knockdown of TgMAP1c reduced the virulence of T. gondii in mice. Taken together, these results demonstrate that TgMAP1c plays a key role in the biogenesis and maintenance of the T. gondii apicoplast.

甲硫氨酸氨肽酶 (MAP) 从新合成的蛋白质中去除 N 端甲硫氨酸，存在于所有生命形式中。弓形虫的基因组中编码了三种 I 型 MAP 和一种 II 型 MAP 。在这项研究中，我们发现每种 I 型Tg MAP（TgMAP1a-c）的诱导型敲低显着降低了寄生虫的生长和增殖。其中，发现Tg MAP1c 定位于寄生虫的顶端。可诱导的Tg敲低MAP1c 导致顶质体编码转录本丰度的缺陷，以及随后顶质体基因组拷贝数的减少和顶质体结构的丢失。这一发现表明顶质体基因组的转录在Tg MAP1c敲低后受损。我们还发现Tg MAP1c 在顶质体生物发生中的功能取决于其酶结构域。Tg MAP1c的活性结构域被Tg MAP1a 或Tg MAP1b的活性结构域取代的重组蛋白的表达不能恢复由敲低Tg引起的生长和复制缺陷表型MAP1c，表明这三种酶具有不同的底物偏好。一项体外分析还表明，Tg MAP1c 是一种活性酶，它专门作用于底物 H-Met- p - NA。此外，Tg MAP1c 的诱导性敲低降低了小鼠体内弓形虫的毒力。总之，这些结果表明Tg MAP1c 在弓形虫顶端的生物发生和维持中起着关键作用。