Delineation of G‐Quadruplex Alkylation Sites Mediated by 3,6‐Bis(1‐methyl‐4‐vinylpyridinium iodide)carbazole‐Aniline Mustard Conjugates |
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Authors: | Chien‐Han Chen Tsung‐Hao Hu Tzu‐Chiao Huang Ying‐Lan Chen Prof. Yet‐Ran Chen Prof. Chien‐Chung Cheng Prof. Chao‐Tsen Chen |
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Affiliation: | 1. Department of Chemistry, National Taiwan University, No. 1, Sec. 4, Roosevelt Road, Taipei, 10617 Taiwan (R.O.C.), Fax: (+886)?2‐23636359;2. Agricultural Biotechnology Research Center, Academia Sinica, No. 128, Sec. 2, Academia Road, Taipei, 11529 Taiwan (R.O.C.);3. Institute of Plant Biology and Department of Life Science, National Taiwan University, No. 1, Sec. 4, Roosevelt Road, Taipei, 10617 Taiwan (R.O.C.);4. Department of Applied Chemistry, Chia‐Yi University, No. 300, Xuefu Road, Chiayi City, 60004 Taiwan (R.O.C.), Fax: (+886)?5‐2717901 |
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Abstract: | A new G‐quadruplex (G‐4)‐directing alkylating agent BMVC‐C3M was designed and synthesized to integrate 3,6‐bis(1‐methyl‐4‐vinylpyridinium iodide)carbazole (BMVC) with aniline mustard. Various telomeric G‐4 structures (hybrid‐2 type and antiparallel) and an oncogene promoter, c‐MYC (parallel), were constructed to react with BMVC‐C3M, yielding 35 % alkylation yield toward G‐4 DNA over other DNA categories (<6 %) and high specificity under competition conditions. Analysis of the intact alkylation adducts by electrospray ionization mass spectroscopy (ESI‐MS) revealed the stepwise DNA alkylation mechanism of aniline mustard for the first time. Furthermore, the monoalkylation sites and intrastrand cross‐linking sites were determined and found to be dependent on G‐4 topology based on the results of footprinting analysis in combination with mass spectroscopic techniques and in silico modeling. The results indicated that BMVC‐C3M preferentially alkylated at A15 (H26), G12 (H24), and G2 (c‐MYC), respectively, as monoalkylated adducts and formed A15–C3M–A21 (H26), G12–C3M–G4 (H24), and G2–C3M–G4/G17 (c‐MYC), respectively, as cross‐linked dialkylated adducts. Collectively, the stability and site‐selective cross‐linking capacity of BMVC‐C3M provides a credible tool for the structural and functional characterization of G‐4 DNAs in biological systems. |
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Keywords: | alkylation DNA recognition DNA structures G‐quadruplexes molecular recognition |
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