BACKGROUND Nucleoside diphosphate kinase (NDK) is a housekeeping enzyme that plays key roles in nucleotide recycling and homeostasis in trypanosomatids. It is also secreted by the intracellular parasite Leishmania to modulate the host response. These functions make NDK an attractive target for drug design and for studies aiming at a better understanding of the mechanisms mediating host-pathogen interactions. RESULTS We report the crystal structure and biophysical characterization of the NDK from Leishmania braziliensis (LbNDK). The subunit consists of six α-helices along with a core of four β-strands arranged in a β2β3β1β4 antiparallel topology order. In contrast to the NDK from L. major, the LbNDK C-terminal extension is partially unfolded. SAXS data showed that LbNDK forms hexamers in solution in the pH range from 7.0 to 4.0, a hydrodynamic behavior conserved in most eukaryotic NDKs. However, DSF assays show that acidification and alkalization decrease the hexamer stability. CONCLUSIONS Our results support that LbNDK remains hexameric in pH conditions akin to that faced by this enzyme when secreted by Leishmania amastigotes in the parasitophorous vacuoles (pH 4.7 to 5.3). The unusual unfolded conformation of LbNDK C-terminus decreases the surface buried in the trimer interface exposing new regions that might be explored for the development of compounds designed to disturb enzyme oligomerization, which may impair the important nucleotide salvage pathway in these parasites.

中文翻译：

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巴西利什曼原虫核苷二磷酸激酶的晶体结构和生物物理特征。


背景技术核苷二磷酸激酶（NDK）是一种管家酶，在锥虫中的核苷酸循环和稳态中发挥关键作用。它也由细胞内寄生虫利什曼原虫分泌来调节宿主反应。这些功能使 NDK 成为药物设计和旨在更好地了解介导宿主与病原体相互作用的机制的研究的有吸引力的目标。结果我们报告了巴西利什曼原虫 (LbNDK) NDK 的晶体结构和生物物理特征。该亚基由六个 α 螺旋和四个 β 链核心组成，这些 β 链以 β2β3β1β4 反平行拓扑顺序排列。与 L. Major 的 NDK 相比，LbNDK C 端延伸部分未折叠。 SAXS 数据显示，LbNDK 在 pH 值范围为 7.0 至 4.0 的溶液中形成六聚体，这是大多数真核 NDK 中保守的流体动力学行为。然而，DSF 测定表明酸化和碱化会降低六聚体的稳定性。结论 我们的结果支持 LbNDK 在类似于利什曼原虫无鞭毛体在寄生液泡中分泌时所面临的 pH 条件下（pH 4.7 至 5.3）仍保持六聚体。 LbNDK C 末端不寻常的未折叠构象减少了埋在三聚体界面中的表面，暴露出新的区域，可用于开发旨在干扰酶寡聚化的化合物，这可能会损害这些寄生虫中重要的核苷酸挽救途径。