The production of the recombinant human MAT2A protein labeled with a 6xHis tag at the C-terminus is initiated by co-cloning the MAT2A gene (1-395aa) with the tag gene into an expression vector. The constructed vectors are introduced into E.coli cells. The transformed E.coli cells are grown under optimal conditions, and IPTG is used to induce expression. The cells are lysed, and the recombinant MAT2A protein is purified using Ni-NTA affinity chromatography, where the His tag binds to nickel ions. Elution is performed using a gradient of imidazole to recover the protein in high purity. SDS-PAGE analysis confirms a purity of over 85% of the recombinant MAT2A protein.
The human MAT2A protein is a crucial enzyme involved in the metabolism of methionine, a sulfur-containing amino acid. MAT2A catalyzes the conversion of L-methionine and ATP into S-adenosylmethionine (SAM), which serves as a universal methyl donor in numerous biological processes, including DNA methylation, RNA methylation, and the synthesis of polyamines and phospholipids [1][2]. This enzyme is particularly significant in the context of cancer, where its expression is often upregulated in various malignancies, including hepatocellular carcinoma (HCC) and breast cancer [3][4].
The expression of MAT2A is typically low in normal adult liver tissues, where MAT1A predominates; however, a switch to MAT2A expression occurs in response to liver injury or during the progression of liver cancer [3][5][6]. This switch is associated with metabolic reprogramming that supports cancer cell proliferation and survival, highlighting MAT2A's role in tumorigenesis [7][8]. Studies have shown that MAT2A expression is induced by hypoxic conditions, which are common in tumor microenvironments, further linking it to cancer progression [5]. The inhibition of MAT2A has been shown to selectively reduce the growth of cancer cells that lack the methylthioadenosine phosphorylase (MTAP) gene, indicating a synthetic lethality approach that could be exploited in cancer therapies [9][10].
References:
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[2] M. Li, Z. Konteatis, N. Nagaraja, Y. Chen, S. Zhou, G. Mae, et al. Leveraging structure-based drug design to identify next-generation mat2a inhibitors, including brain-penetrant and peripherally efficacious leads, Journal of Medicinal Chemistry, vol. 65, no. 6, p. 4600-4615, 2022. https://doi.org/10.1021/acs.jmedchem.1c01595
[3] Q. Liu, J. Chen, L. Liu, J. Zhang, D. Wang, L. Ma, et al. The x protein of hepatitis b virus inhibits apoptosis in hepatoma cells through enhancing the methionine adenosyltransferase 2a gene expression and reducing s-adenosylmethionine production, Journal of Biological Chemistry, vol. 286, no. 19, p. 17168-17180, 2011. https://doi.org/10.1074/jbc.m110.167783
[4] K. Secker, B. Bloechl, H. Keppeler, S. Duerr-Stoerzer, H. Schmid, D. Schneidawind, et al. Mat2a as key regulator and therapeutic target in mllr leukemogenesis, Cancers, vol. 12, no. 5, p. 1342, 2020. https://doi.org/10.3390/cancers12051342
[5] Q. Liu, L. Liu, Y. Zhao, J. Zhang, D. Wang, J. Chen, et al. Hypoxia induces genomic dna demethylation through the activation of hif-1α and transcriptional upregulation of mat2a in hepatoma cells, Molecular Cancer Therapeutics, vol. 10, no. 6, p. 1113-1123, 2011. https://doi.org/10.1158/1535-7163.mct-10-1010
[6] C. Fusco, M. Schimpl, U. Börjesson, T. Cheung, I. Collie, L. Evans, et al. Fragment-based design of a potent mat2a inhibitor and in vivo evaluation in an mtap null xenograft model, Journal of Medicinal Chemistry, vol. 64, no. 10, p. 6814-6826, 2021. https://doi.org/10.1021/acs.jmedchem.1c00067
[7] E. Strekalova, D. Malin, E. Weisenhorn, J. Russell, D. Hoelper, A. Jainet al., S-adenosylmethionine biosynthesis is a targetable metabolic vulnerability of cancer stem cells, Breast Cancer Research and Treatment, vol. 175, no. 1, p. 39-50, 2019. https://doi.org/10.1007/s10549-019-05146-7
[8] M. Simile, G. Peitta, M. Tomasi, S. Brozzetti, C. Feo, A. Porcu, et al. Microrna-203 impacts on the growth, aggressiveness and prognosis of hepatocellular carcinoma by targeting mat2a and mat2b genes, Oncotarget, vol. 10, no. 29, p. 2835-2854, 2019. https://doi.org/10.18632/oncotarget.26838
[9] P. Kalev, M. Hyer, S. Größ, Z. Konteatis, C. Chen, M. Fletcher, et al. Mat2a inhibition blocks the growth of mtap-deleted cancer cells by reducing prmt5-dependent mrna splicing and inducing dna damage, Cancer Cell, vol. 39, no. 2, p. 209-224.e11, 2021. https://doi.org/10.1016/j.ccell.2020.12.010
[10] Z. Lou, Targeting mtap creates a therapeutic vulnerability to parp inhibition in brain metastatic tnbc by disrupting mat2a mediated methionine metabolism,, 2023. https://doi.org/10.21203/rs.3.rs-3579438/v1
