2025-06-18 イェール大学
<関連情報>
- https://medicine.yale.edu/news-article/how-parasitesand-a-bit-of-serendipityled-to-a-powerful-new-drug-discovery-tool-a-qanda-with-choukri-ben-mamoun/
- https://www.science.org/doi/10.1126/sciadv.adv2397
- https://www.sciencedirect.com/science/article/abs/pii/S0076687925000953
- https://www.jbc.org/article/S0021-9258(24)02334-2/fulltext
スペルミジンは、細胞内寄生虫が宿主赤血球内で生存するために必要な重要なポリアミンであるSpermidine is a key polyamine required by intracellular parasites for survival within host erythrocytes
Pallavi Singh, Jae-Yeon Choi, Emmanuel Cornillot, and Choukri Ben Mamoun
Science Advances Published:18 Jun 2025
DOI:https://doi.org/10.1126/sciadv.adv2397
Abstract
Intracellular parasites, including Babesia and Plasmodium, the agents of human babesiosis and malaria, depend on the salvage or de novo synthesis of critical nutrients for survival within human erythrocytes. Among these, polyamines play a pivotal role, but their specific requirements and molecular functions in intraerythrocytic parasites remain poorly understood. We identify spermidine as a key polyamine for Babesia duncani and Plasmodium falciparum for intraerythrocytic development. We demonstrate that spermidine is indispensable for regulating protein translation through hypusination of the eukaryotic translation initiation factor eIF5A, and its depletion leads to increased production of reactive oxygen species. Disruption of spermidine biosynthesis or its conversion from spermine results in parasite death. We also show that B. duncani and other Babesia species use an ancestral spermidine synthase–like enzyme, highlighting a distinct evolutionary adaptation from P. falciparum. Our results reveal the spermidine’s dual role in oxidative stress defense and translation regulation, positioning spermidine biosynthesis as a critical vulnerability and a promising therapeutic target.
第21章-アミノプロピルトランスフェラーゼ活性を測定するための蛍光アッセイ Chapter Twenty One – A fluorescence-based assay for measuring aminopropyltransferase activity
Pallavi Singh, Jae-Yeon Choi, Choukri Ben Mamoun
Methods in Enzymology Available online: 11 March 2025
DOI:https://doi.org/10.1016/bs.mie.2025.01.067
Abstract
Polyamines (PAs) are small polycationic alkylamines that are essential for numerous cellular processes and found in all living cells. The three principal polyamines, putrescine (PUT), spermidine (SPD), and spermine (SPM), have been shown to play crucial roles in cellular function and implicated in several diseases including infectious diseases, cancer and neurodegenerative disorders. As such, the enzymes involved in polyamine biosynthesis are promising targets for developing antimicrobial, antineoplastic and neuroprotective therapies. Aminopropyl transferases (APTs) are key enzymes in this pathway, catalyzing the formation of spermidine from putrescine and spermine from spermidine. While in most eukaryotes and prokaryotes, the spermidine synthase and spermine synthase activities are catalyzed by distinct enzymes, some organisms such as Plasmodium falciparum have a single enzyme, which catalyzes both reactions with varying efficiency. To date, efforts to inhibit APTs have focused primarily on substrate analogs, often with limited selectivity. A major challenge in discovering novel inhibitors has been the lack of an assay suitable for high-throughput chemical screening. We have recently developed DAB-APT, the first fluorescence-based assay for measuring APT activity, using 1,2-diacetyl benzene (DAB) which reacts with putrescine, spermidine, and spermine to form fluorescent conjugates, with fluorescence intensity correlating to carbon chain length. The DAB-APT assay has been validated using APT enzymes from Saccharomyces cerevisiae, and P. falciparum, and has been found to be suitable for high-throughput screening of large chemical libraries. This assay represents a significant advancement, offering a valuable tool for identifying potential inhibitors of APT enzymes and accelerating drug discovery efforts in cancer, neurobiology, and infectious diseases.
ポリアミン生合成アミノプロピルトランスフェラーゼを介した触媒反応を測定する蛍光ベースのアッセイ法 A fluorescence-based assay for measuring polyamine biosynthesis aminopropyl transferase–mediated catalysis
Pallavi Singh ∙ Jae-Yeon Choi ∙ Weiwei Wang ∙ … ∙ Sovitj Pou, ∙ Aaron Nilsen ∙ Choukri Ben Mamoun
Journal of Biological Chemistry Published:September 27, 2024
DOI:https://doi.org/10.1016/j.jbc.2024.107832
Abstract
Polyamines are polycationic molecules that are crucial in a wide array of cellular functions. Their biosynthesis is mediated by aminopropyl transferases (APTs), which are promising targets for antimicrobial, antineoplastic, and antineurodegenerative therapies. A major limitation in studying APT enzymes, however, is the lack of high-throughput assays to measure their activity. We have developed the first fluorescence-based assay, diacetyl benzene (DAB)-APT, for the measurement of APT activity using 1,2-DAB, which forms fluorescent conjugates with putrescine, spermidine, and spermine, with fluorescence intensity increasing with the carbon chain length. The assay has been validated using APT enzymes from Saccharomyces cerevisiae and Plasmodium falciparum, and the data further validated by mass spectrometry and TLC. Using mass spectrometry analysis, the structures of the fluorescent putrescine, spermidine, and spermine 1,2-DAB adducts were determined to be substituted 1,3-dimethyl isoindoles. The DAB-APT assay is optimized for high-throughput screening, facilitating the evaluation of large chemical libraries. Given the critical roles of APTs in infectious diseases, oncology, and neurobiology, the DAB-APT assay offers a powerful tool with broad applicability, poised to drive advancements in research and drug discovery.
