Supporting Information Copyright Wiley-VCH Verlag GmbH & Co. KGa, 6945 Weinheim, 28
Label-Free Fluorescent Probing of G-Quadruplex Formation and Real-Time Monitoring of DN Folding by a Quaternized Tetraphenylethene Salt with ggregation-induced Emission Characteristics Yuning Hong, [a] Matthias Häuβler, [a] Jacky W. Y. Lam, [a] Zhen Li, [a] King Keung Sin, [a] Yongqiang Dong, [a, b] Hui Tong, [a] Jianzhao Liu, [a] njun Qin, [a, b] Reinhard Renneberg, [a] [a, b] and en Zhong Tang* [a]department of Chemistry The Hong Kong University of Science & Technology Clear Water ay, Kowloon, Hong Kong (China) [b]department of Polymer Science and Engineering Zhejiang University Hangzhou 327 (China)
Supporting Information for chem.27723 Experimental Section General Information: THF (Labscan) was purified by distillation from sodium benzophenone ketyl under nitrogen immediately prior to use. DHP, titanium(iv) chloride, zinc dust,,2-dibromoethane, potassium carbonate, acetone, triethylamine, and other reagents were all purchased from ldrich and used as received. H and 3 C NMR spectra were measured on a uker RX 3 spectrometer with tetramethylsilane (TMS; δ = ) as the internal standard. Mass spectra were recorded on a Finnigan TSQ 7 triple quadrupole spectrometer operating in a MLDI-TF mode. UV spectra were measured on a Milton Roy Spectronic 3 rray spectrophotometer and FL spectra were recorded on a Perkin-Elmer LS 55 spectrofluorometer with a Xenon discharge lamp excitation. Time-dependent FL signals were measured using a Fluostarptima multifunctional microplate reader (MG Labtechnologies) with excitation/emission wavelengths set at 35/47 nm. CD spectra were recorded on a Jasco J-8 spectropolarimeter in mm quartz cuvette using a step resolution of.2 nm, a scan speed of nm/min, a sensitivity of., and a response time of.5 s. Each spectrum was the average of three scans. Dye Synthesis: The synthetic route to TTPE is shown in Scheme S. McMurry coupling of EP yields TEPE, quaternization of which by triethylamine generates TTPE. Detailed experimental procedures for the dye synthesis are given below. Preparation of EP: To a mixture of DHP (3. g, 4. mmol) and potassium carbonate (5. g, 36.2 mmol) in acetone (5 ml) was added,2-dibromoethane (4 ml, 46.4 mmol). The mixture was refluxed under stirring for 24 h. fter filtration and solvent evaporation, the crude product was purified by a silica gel column using chloroform as eluent. EP was obtained as white powder in 7% yield (4.2 g). R f =.6 (chloroform); H NMR (3 MHz, CDCl 3, 25 o C, TMS): δ = 3.68 (t, 4H; CH 2 ), 4.38 (t, 4H; CH 2 ), 6.97 (d, 4H, J 9. Hz; r), 7.78 ppm (d, 4H, J = 9. Hz; r); 3 C NMR (75 MHz, CDCl 3, 25 o C, TMS): δ = 29.3, 68.6, 4.8, 5.8, 3.9, 33., 62. ppm. Synthesis of TEPE: In a suspension of EP (. g, 2.3 mmol) in 5 ml of THF were added TiCl 4 (.26 ml, 2.3 mmol) and Zn dust (.3 g, 4.6 mmol). fter refluxing for 2 h, the reaction mixture was cooled to room temperature and filtered. The solvent was evaporated under vacuum and the crude product was purified by a silica gel column using a chloroform/hexane (:4 v/v) mixture as eluent. TEPE was obtained as white solid in 63% yield (.66 g). R f =.7 (chloroform/hexane = :4); H NMR (3 MHz, CDCl 3, 25 o C, TMS): δ = 3.63 (t, 8H; CH 2 ), 4.23 (t, 8H; CH 2 ), 6.66 (d, 8H, J = 8.7 Hz; r), 6.93 ppm (d, 4H, J = 8.7 Hz; r); 3 C NMR (75 MHz, CDCl 3, 25 o C, TMS): δ = 29.8, 68.3, 4.5, 33.3, 38., 39., 57. ppm; MLDI-TF-MS m/z: calcd for C 34 H 32 4 4 + : 823.8993; found 823.8688 ([M] + ). Synthesis of TTPE: In a 25 ml flask with a magnetic stirrer was dissolved TEPE ( mg,.2 mmol) in THF ( ml). fter adding an excess amount of triethylamine (5 ml, 35.6 mmol), the solution was refluxed for 3 days. During the period, ml of water was added at several intervals. The organic solvents were evaporated under reduced pressure and the aqueous solution was washed with chloroform three times. fter solvent evaporation and drying overnight in vacuo at 5 ºC, TTPE was isolated as yellow viscous liquid in 56% yield (.89 g). H NMR (4 MHz, D 2, 25 o C, TMS): δ =.3.2 (m, 36H; NCH 2 CH 3 ), 3.25 3.3 (m, 24H; NCH 2 CH 3 ), 3.45 3.54 (m, 8H; CH 2 CH 2 N), 4.23 4.24 (m, 8H; CH 2 CH 2 N), 6.65 6.67 (m, 8H; r), 6.89 6.94 ppm (m, 8H; r); 3 C NMR (75 MHz, D 2, 25 o C, TMS): δ = 7.4, 47.2, 54.2, 6.8, 4. 5, 33., 38., 39.5, 56.2 ppm; MLDI-TF-MS m/z: calcd for C 58 H 92 4 N 4 4 + [M 2] + : 66.5485; found 66.5359 ([M 2] + ). [a] [b] [b] Y. ((Title(s), Hong, Initial(s), Dr. M. Häuβler, Surname(s) Dr. of J. uthor(s) W. Y. Lam, including Dr. Corresponding K. K. Sin, Prof. uthor(s))) Dr. Z. Li, Dr. Y. Dong, Dr. H. Tong, J. Liu, Dr.. Qin, Prof. Dr. ((Department)) R. Renneberg, Prof. Dr.. Z. Tang ((Institution)) Department of Chemistry ((ddress The Hong )) Kong University of Science & Technology Fax: Clear (+) Water ay, Kowloon, Hong Kong (China) E-mail: Fax: (+852)2358-594 E-mail: tangbenz@ust.hk ((Title(s), Initial(s), Surname(s) of uthor(s))) ((Department)) Dr. Y. Dong, Dr.. Qin, Prof. Dr.. Z. Tang ((Institution)) Department of Polymer Science and Engineering ((ddress Zhejiang University 2)) Hangzhou 327 (China) ((If supporting information is submitted, please include the following line: Supporting information for this article is available on the WWW under http://www.chemeurj.org/ or from the author.)) CH 2 CH 2, K 2 C 3 H H (CH 3 ) 2 C, reflux DHP EP TEPE Et 3 N, THF/H 2 reflux TTPE TiCl 4, Zn THF, reflux Scheme S. Synthesis of,,2,2-tetrakis[4-(2-triethylammonioethoxy)phenyl]ethene tetrabromide (TTPE).
2 TTPE 6 4 2 8 6 4 2 Chemical shift (ppm) * TEPE Figure S. 3 C NMR spectra of () TEPE in chloroform-d and () TTPE in waterd 2. The solvent peak is marked with an asterisk. bsorbance (au) Water:n (v/v) bsorption 99: : x2.5 25 35 45 55 65 Emission Figure S2. bsorption and emission spectra of TEPE (5 µm) in acetonitrile (n) and an n/water mixture (:99 v/v). - G/Na + G/TTPE G/Ca 2+ /TTPE -2 235 255 275 295 35 Figure S4. CD spectra of G in a Tris HCl buffer in the presence or absence of a metal ion and/or TTPE at 2 ºC. [G] = 9 µm, [ion] =.5 M, [TTPE] = 4.5 µm. 38 43 48 53 58 63 2 3 4 5 K + concn (mm) 5 5 I/I (%) 35 28 2 4 7 Wavelength Intensity K + concentration (mm) Figure S5. () Emission spectra of TTPE in a Tris HCl buffer in the presence of G and K +. () Effects of [K + ] on emission intensity at 47 nm and peak wavelength (λ em ) of the TTPE/G solution. [TTPE] = 4.5 µm, [G] = 9 µm; λ ex = 35 nm. 25 2 5 5 K + added to TTPE/G G added to TTPE/K + TTPE added to G/K + 38 45 52 59 66 3.6 2.7.8.9. 49 48 47 46 λ em (nm) -.9 235 255 275 295 35 K + added to TTPE/G G added to TTPE/K + TTPE added to G/K + TTPE concn (µm) 22.7 45.4 8.2 3.6 4.5.9. Figure S6. Effects of addition sequence on () FL and () CD spectra of K + /TTPE/ G complex in 5 mm Tris HCl buffer solutions. [K + ] =.5 M, [TTPE] = 4.5 µm, [G] = 9 µm; λ ex = 35 nm. TTPE/G/Li + TTPE/G/Ca 2+ TTPE/G/NH + 4 TTPE/G/Mg 2+ TTPE/Ca 2+ TTPE/Li + TTPE/Mg 2+ TTPE/Na + TTPE/NH + 4 38 43 48 53 58 63 Figure S3. Emission spectra of TTPE in the presence of a solution of G (4.5 µm) in 5 mm Tris HCl buffer (ph = 7.5). 38 43 48 53 58 38 43 48 53 58 Figure S7. FL spectra of () TTPE/G and () TTPE in a Tris HCl buffer in the presence of cationic species. [TTPE] = 4.5 µm, [G] = 9 µm, [ion] =.5 M; λ ex = 35 nm. 2
2 - -2 K + /Na + : : 3 : 6 :2 235 255 275 295 35 TTPE/G/C2 TTPE/G/C2/Na + TTPE/G/C2/Ca 2+ 38 43 48 53 58 63 Figure S8. CD spectra of the G/TTPE/K + solutions in a Tris HCl buffer titrated by different amounts of Na + ions. [G] = 9 µm, [TTPE] = 4.5 µm, [K + ] = mm; λ ex = 35 nm. Figure S. Emission spectra of buffer solutions of TTPE/C2 in the absence and presence of metal ions. [TTPE] = 4.5 µm, [G] = [C2] = 4.5 µm, [ion] =.5 M; λ ex = 35 nm. TTPE/C TTPE/C/Li + TTPE/C/Na + TTPE/C/K + TTPE/C/Ca 2+ TTPE/G/C TTPE/G/C/K + TTPE/G/C/Na + TTPE/G/C/Ca 2+ K + Time (s) 72 6 5 42 3 8 2 6 Na + NH + 4 Ca 2+ 38 43 48 53 58 63 38 43 48 53 58 63 Figure S9. Emission spectra of buffer solutions (ph = 7.5) of () TTPE/C and () TTPE/G/C in the absence and presence of metal ions. [TTPE] = 4.5 µm, [ion] =.5 M; λ ex = 35 nm; [C] = 9 µm (for ), [G] = [C] = 4.5 µm (for ). 38 43 48 53 58 63 3 6 9 2 5 Time (min) Figure S2. Time courses of evolution of emission intensities of buffer solutions of TTPE/G after addition of cationic species. [TTPE] = 4.5 µm, [G] = 9 µm; λ ex = 35 nm; for : [K + ] =.5 M; for : [Na + ] = [NH 4 + ] =.5 M, [Ca 2+ ] =.25 M..6.3. -.3 G/C G/C/K + -.6 25 24 265 29 35 Figure S3. () Selected crystal structure of G-quadruplex of a human telomeric DN (data taken from RSC Protein Data ank; ID No. KF) and () molecular structure of TTPE with minimized energy, simulated by molecular mechanics MM2 program installed in Chem3D Ultra 8.. Figure S. CD spectra of G/C in the absence or presence of K + after hybridization in a Tris HCl buffer (ph = 7.5). [G] = [C] = 4.5 µm, [K + ] =.5 M; λ ex = 35 nm. 3