Supporting Information

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1 Supporting Information Tuning Supramolecular Structure and Functions of Peptide bola-amphiphile by Solvent Evaporation-Dissolution Anhe Wang,, Lingyun Cui,, Sisir Debnath, Qianqian Dong, Xuehai Yan, Xi Zhang, Rein V. Ulijn *,,ǁ,, and Shuo Bai *, State Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing, China. WestCHEM, Department of Pure and Applied Chemistry, University of Strathclyde, Glasgow G1 1XL, U.K. MOE Key Lab of Organic Optoelectronics & Molecular Engineering, Department of Chemistry, Tsinghua University, Beijing, China. ǁ Advanced Science Research Center (ASRC), City University of New York, New York, NY10031, USA. Hunter College, Department of Chemistry and Biochemistry, 695 Park Avenue, New York, New York 10065, USA. The Graduate Center of the City University of New York, New York 10016, USA. These authors contributed equally to this work. * S-1

2 Synthesis of PBI-[GY] 2 Scheme S1: Synthesis route of PBI-[GY] 2. Synthesis of PBI-[GY] 2 : Boc-glycine (1 g, 4.64 mmol), L-tyrosine tert-butyl ester ( g, 4.21 mmol) and HBTU (1.91g, 5.02 mmol) were mixed together in 10 ml dry DMF. Then 1.93 ml (10.52 mmol) of DIPEA was added to this solution and the mixture was stirred overnight under nitrogen atmosphere. After reaction, the product was extracted by 75 ml of ethyl acetate after successive wash with 25 ml of 1 N NaHCO 3 and 25 ml of 1 N hydrochloric acid, and then dried over MgSO 4. After evaporation of the solvent, the compound was purified by column chromatography on silica gel using dichloromethane/ methanol (96:4) as eluent (1.3 g, 71 %). After purification, the Boc group of N-terminus and tert-butyl group of C-terminus was removed by one step reaction using trifluoroacetic acid (TFA, 5 ml) in dry dichloromethane for 24 hours. The excess TFA was removed by high vacuum pump and the product was washed thoroughly for 3 times with diethyl ether to get pure GY dipeptide (0.76 gm, 97%). Finally, the dipeptide was attached at the both ends of perylene-3,4,9,10-tetracarboxylic dianhydride. 0.3g (0.76 mmol) of Perylene-3,4,9,10-tetracarboxylic dianhydride was mixed S-2

3 with 0.73 g (3.06 mmol) of GY dipeptide, 10 g of imidazole as a solvent and g (0.79 mmol) of Zinc acetate. The mixture was heated at 130 o C for 24 hours under nitrogen atmosphere. The product was cooled down and imidazole dissolved in 1 N hydrochloride acid filtered in the product. The residue was collected and dissolved in 1 N NaOH, filtered, and precipitated out by adding 1 N HCl. This process was repeated for four times to give pure PBI-[GY] 2. The final yield of the product was 5 g. 1 H NMR (DMSO-d 6, 90 o C, 400 MHz) δ (m, 4H, CH 2 in tyrosine moiety), (m, 2H, CH in tyrosine moiety), 4.74 (s, 4H, N-CH 2 ), 6.68 (d, 4H, CH at ortho position of OH group of tyrosine, J = 8.8 Hz), 7.03 (d, 4H, CH at meta position of OH group of tyrosine, J = 8.8 Hz), 8.19 (d, 2H, CONH, J = 7.4 Hz), (m, 4H, perylene aromatic CH), (m, 4H, perylene aromatic CH). 13 C NMR (DMSO, 100 MHz): δ 35.8 (Tyr CH 2 ), 47.6 (NCH 2 ), 53.4 (Tyr chiral CH), (Ar CH), (Ar CH), (Ar CH), (Ar C q ), (ArC q ), (Ar CH), (ArC q ), (ArC q ), (Ar C q ), (ArC q ), 17 (C=O), (C=O), (C=O). ESI-MS: m/z: calculated for C 46 H 32 N 4 O 12 : ; found: 83 [M -H]. S-3

4 Figure S1. TEM (left) and AFM (right) images of structures of PBI-[GY] 2 assembled in THF. The concentration is 1x10-4 M. Scale bar of TEM and AFM images are 100 nm and 500 nm, respectively. Figure S2. Size distribution of dynamic light scattering measurement of co-assembled nanostructures of PBI-[GY] 2 with trace THF in buffer solution via solvent-evaporation procedure. The concentration is 1x10-4 M. S-4

5 00 PBI-[GY] 2 /trace THF in water PBI-[GY] 2 in water Transmittance (%) Wavenumber (cm -1 ) Figure S3. FTIR spectra of PBI-[GY] 2 in water without (red curve) or with (black curve) trace THF. The concentration is 1x10-4 M. Normalized Absorption UV-Vis spectrum Fluorescence spectrum THF Normalized Intensity Figure S4. UV-Vis (black curve) and fluorescence emission spectra (red curve) of PBI-[GY] 2 in THF. The concentration is 1x10-4 M. Insert is optical photograph of PBI-[GY] 2 THF solution at the same concentration under UV lamp (365 nm). S-5

6 a) 0.1%THF in water 1%THF in water 10%THF in water b) Normalized Absorption Water 0.1% THF in water 1% THF in water 10% THF in water Figure S5. a) TEM images of PBI-[GY] 2 assembled structures in mixed buffer solution with varied amount of THF (The weight fractions of THF in water are 0.1, 1 and 10%.) The concentration is 1x10-4 M. Scale bar of TEM images is 200 nm. b) UV-Vis (left) and fluorescence emission spectra (right) of PBI-[GY] 2 in mixed buffer solution with varied amount of THF (The weight fractions of THF in water are 0.1, 1 and 10%.) The concentration is 1x10-4 M. The excitation wavelength is 365 nm. Intensity (a.u.) Water 0.1% THF in water 1% THF in water 10% THF in water a) PBI-[GY] 2 in ethanol PBI-[GY] 2 /trace ethanol in water b) Absorption 1.2 PBI-[GY] 2 in water PBI-[GY] 2 /EtOH in water Figure S6. a) TEM images of structures of PBI-[GY] 2 assembled in ethanol (left) and in buffer solution co-assembled with trace ethanol via evaporation-dissolution treatment. The concentration is 1x10-4 M. Scale bar of TEM images is 200 nm. b) UV-Vis (left) and fluorescence emission spectra (right) of PBI-[GY] 2 with (red curve) or without (black curve) trace ethanol in buffer solution. The concentration is 1x10-4 M. The excitation wavelength is 365 nm. S-6 Intensity (a.u.) PBI-[GY] 2 in water PBI-[GY] 2 /EtOH in water

7 Figure S7. a) In vitro viability of 3T3 cells by CCK-8 method in the presence of PBI-[GY] 2 aggregates with or without trace THF. The excitation wavelength is 490 nm. Cell staining experiments by using PBI-[GY] 2 aggregates self-assembled without or with trace THF. CLSM fluorescent images: the corresponding overlapped fluorescence images of 3T3 cells incubated with PBI-[GY] 2 aggregates without b) and with trace THF c) for 12 h, respectively. The concentration of PBI-[GY] 2 was 4 mg ml -1. The nuclei and membrane of 3T3 cells were stained by Hoechst (blue fluorescent domains) and Alexa Fluor 488 (green ones), respectively. S-7