PDF(Pubmed)
Abstract：
Plasmodium falciparum is the main causative agent of malaria, a deadly disease that mainly affects children under five years old. Artemisinin-based combination therapies have been pivotal in controlling the disease, but resistance has arisen in various regions, increasing the risk of treatment failure. The non-mevalonate pathway is essential for the isoprenoid synthesis in Plasmodium and provides several under-explored targets to be used in the discovery of new antimalarials. 1-deoxy-D-xylulose-5-phosphate synthase (DXPS) is the first and rate-limiting enzyme of the pathway. Despite its importance, there are no structures available for any Plasmodium spp., due to the complex sequence which contains large regions of high disorder, making crystallisation a difficult task. In this manuscript, we use cryo-electron microscopy to solve the P. falciparum DXPS structure at a final resolution of 2.42 Å. Overall, the structure resembles other DXPS enzymes but includes a distinct N-terminal domain exclusive to the Plasmodium genus. Mutational studies show that destabilization of the cap domain interface negatively impacts protein stability and activity. Additionally, a density for the co-factor thiamine diphosphate is found in the active site. Our work highlights the potential of cryo-EM to obtain structures of P. falciparum proteins that are unfeasible by means of crystallography.
摘要：
恶性疟原虫是疟疾的主要病原体，一种主要影响五岁以下儿童的致命疾病。以青蒿素为基础的联合疗法在控制疾病方面至关重要，但是不同地区出现了抵抗，增加治疗失败的风险。非甲羟戊酸途径对于疟原虫中的类异戊二烯合成至关重要，并提供了一些尚未开发的靶标，用于发现新的抗疟疾药。1-脱氧-D-木酮糖-5-磷酸合酶（DXPS）是该途径的第一个和限速酶。尽管它很重要,任何疟原虫都没有可用的结构。,由于复杂的序列包含大量高度无序的区域，使结晶成为一项艰巨的任务。在这份手稿中,我们使用低温电子显微镜来解决恶性疟原虫DXPS结构，最终分辨率为2.42µ。总的来说,该结构类似于其他DXPS酶，但包括疟原虫属独有的独特N末端结构域。突变研究表明，帽结构域界面的不稳定对蛋白质的稳定性和活性产生负面影响。此外,在活性位点发现辅因子硫胺素二磷酸酯的密度。我们的工作强调了冷冻EM获得恶性疟原虫蛋白质结构的潜力，这些结构通过晶体学是不可行的。
