1Department of Medical Environmental Biology and Tropical Medicine, School of Medicine, Kangwon National University, Chuncheon 24341, Korea 2Department of Epidemiology and Tropical Diseases, Faculty of Public Health, Universitas Diponegoro, Semarang 50275, Indonesia 3Department of Biochemistry, School of Medicine, Inha University, Incheon 22212, Korea 4Department of Biomedical Science, School of Medicine, Inha University, Incheon 22212, Korea 5Institute of Medical Sciences, Kangwon National University, Chuncheon 24341, Korea 6Department of Parasitology and Tropical Medicine, School of Medicine, Inha University, Incheon 22212, Korea
Clonorchis sinensis is a liver fluke that causes clonorchiasis, a significant public health concern in East Asia, closely associated with hepatobiliary diseases. Dopamine is an essential neurotransmitter involved in neuromuscular signaling, and its uptake by trematodes may contribute to parasite physiology and survival. This study aimed to characterize the dopamine transporter CsDAT in C. sinensis by synthesizing cDNA from adult worms and expressing it in Xenopus laevis oocytes; subsequently, uptake assays were conducted using radiolabeled dopamine. Functional assays confirmed that CsDAT mediates dopamine uptake in a sodium-dependent manner. The uptake was saturable and exhibited Michaelis-Menten kinetics with a Michaelis constant of 454.5 nM and a maximum uptake rate of 1,422.5 fmol/oocyte/h. CsDAT efficiently transported dopamine with high affinity, indicating its physiological relevance in the parasite. A 3-dimensional model of CsDAT was constructed to examine its structural features. The predicted structure contained a conserved substrate-binding pocket similar to that of other known neurotransmitter transporters. Molecular docking simulations showed that dopamine stably fits within the binding pocket. The key amino acid residues formed hydrogen bonds and hydrophobic interactions with dopamine. Interestingly, dopamine and several inhibitors demonstrated higher binding affinity to CsDAT than the human dopamine transporter. This study provides the first functional and structural insights into CsDAT. The higher inhibitor-binding affinity of CsDAT compared to human dopamine transporter suggests its potential for use in therapeutic exploration. Targeting CsDAT may facilitate the development of new therapeutic agents against clonorchiasis with minimal off-target effects on the human nervous system.
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