The generation of recombinant human IDO1 protein starts with cloning the IDO1 gene (1-403aa) with the N-terminal 6xHis-tag gene into an expression vector, followed by its introduction into E.coli cells. Once expressed, the protein is isolated and purified using affinity chromatography from the cell lysate. Following purification, SDS-PAGE is employed to determine the purity of the recombinant IDO1 protein, exceeding 90%. This ensures the protein's suitability for applications in fields like cardiovascular research.
Human IDO1 is a crucial enzyme involved in the catabolism of the essential amino acid tryptophan (Trp) through the kynurenine pathway. It catalyzes the oxidative cleavage of Trp to N-formyl kynurenine, which subsequently hydrolyzes to kynurenine (Kyn) [1][2]. This reaction represents the rate-limiting step in the kynurenine pathway, which is significant for various physiological processes, including immune regulation and metabolism [3][4]. IDO1 is primarily expressed in multiple tissues, including lymphoid organs and the placenta, and its activity is induced by pro-inflammatory cytokines such as interferon-gamma (IFN-γ) [5][6].
By degrading Trp, IDO1 creates a local environment unfavorable for T-cell proliferation and function, leading to immunosuppression [7][8], which is exploited by tumors to evade immune detection, as the depletion of Trp and the accumulation of Kyn can inhibit T-cell activity and promote tumor growth [1][9]. Furthermore, high levels of IDO1 expression have been associated with poor clinical outcomes in various cancers, indicating its potential as a therapeutic target [9][4].
References:
[1] P. Azimnasab-Sorkhabi, M. Soltani-Asl, T. Yoshinaga, M. Dagli, C. Massoco, & J. Kfoury. Indoleamine-2,3 dioxygenase: a fate-changer of the tumor microenvironment, Molecular Biology Reports, vol. 50, no. 7, p. 6133-6145, 2023. https://doi.org/10.1007/s11033-023-08469-3
[2] A. Lewis-Ballester, S. Karkashon, D. Batabyal, T. Poulos, & S. Yeh. Inhibition mechanisms of human indoleamine 2,3 dioxygenase 1, Journal of the American Chemical Society, vol. 140, no. 27, p. 8518-8525, 2018. https://doi.org/10.1021/jacs.8b03691
[3] S. Oda, H. Sugimoto, T. Yoshida, & Y. Shiro. Crystallization and preliminary crystallographic studies of human indoleamine 2,3-dioxygenase, Acta Crystallographica Section F Structural Biology and Crystallization Communications, vol. 62, no. 3, p. 221-223, 2006. https://doi.org/10.1107/s1744309106003356
[4] U. Röhrig, S. Majjigapu, P. Vogel, V. Zoete, & O. Michielin, Challenges in the discovery of indoleamine 2,3-dioxygenase 1 (ido1) inhibitors, Journal of Medicinal Chemistry, vol. 58, no. 24, p. 9421-9437, 2015. https://doi.org/10.1021/acs.jmedchem.5b00326
[5] P. Terness, T. Bauer, et al. Inhibition of allogeneic t cell proliferation by indoleamine 2,3-dioxygenase–expressing dendritic cells, The Journal of Experimental Medicine, vol. 196, no. 4, p. 447-457, 2002. https://doi.org/10.1084/jem.20020052
[6] Y. Kudo, I. Koh, & J. Sugimoto. Localization of indoleamine 2,3-dioxygenase-1 and indoleamine 2,3-dioxygenase-2 at the human maternal-fetal interface, International Journal of Tryptophan Research, vol. 13, p. 117864692098416, 2020. https://doi.org/10.1177/1178646920984163
[7] Y. Kudo. The role of placental indoleamine 2,3-dioxygenase in human pregnancy, Obstetrics & Gynecology Science, vol. 56, no. 4, p. 209, 2013. https://doi.org/10.5468/ogs.2013.56.4.209
[8] X. Song, Q. Si, et al. Indoleamine 2,3-dioxygenase 1: a promising therapeutic target in malignant tumor, Frontiers in Immunology, vol. 12, 2021. https://doi.org/10.3389/fimmu.2021.800630
[9] M. Chamuleau, A. Loosdrecht, et al. High indo (indoleamine 2,3-dioxygenase) mrna level in blasts of acute myeloid leukemic patients predicts poor clinical outcome, Haematologica, vol. 93, no. 12, p. 1894-1898, 2008. https://doi.org/10.3324/haematol.13113
