SupplementaryInformation

Synthesis and protein binding properties of novel highly functionalized Calix[4]arene

Reza Zadmard*, ShukufeDarvishi, Peyman Akbari-Moghaddam

Chemistry and Chemical Engineering Research Center of Iran (CCERCI), PO Box 14335-186, Tehran, Iran,

E-mail:

Table of contents / Page
Figure S1. 1H NMR (400 MHz, DMSO-d6) spectrum of compound5 / 3
Figure S2. 13C NMR (100 MHz, DMSO-d6) spectrum of compound5 / 3
Figure S3. FT-IR spectrum of compound5 / 4
Figure S4. High resolution mass spectrum (HRMS) ofcompound5 / 4
Fluorescence measurements / 5
Table S1. Intrinsic fluorescence of Lys upon interaction with compound 5 / 6
Table S2. Intrinsic fluorescence of BLG upon interaction with compound 5 / 7
Table S3. Intrinsic fluorescence of BSA upon interaction with compound 5 / 8
Table S4. Intrinsic fluorescence of HSA upon interaction with compound 5 / 9

Figure S1. 1H NMR (400 MHz, DMSO-d6) spectrum of compound5

Figure S2. 13C NMR (100 MHz, DMSO-d6) spectrum of compound5

Figure S3. FT-IR spectrum of compound5

Figure S4. High resolution mass spectrum (HRMS) of compound 5

Fluorescence measurements

Fluorescent spectra were recorded on aJASCO FP-6500 spectrofluorometer. Protein solution (1 µM, prepared in a suitable buffer immediately before the experiment) was prepared in a 1-cm path length fluorescence cuvette.Calix[4]arene5 solution (10 µM) was consecutively added in μLvolumes using trace Eppendorf pipette to the fluorescence cuvette. Protein intrinsic fluorescence was recorded from 300 to 500 nm using an excitation wavelength of 280 nm at 25 ºC.

Table S1. Intrinsic fluorescence of Lys upon interaction with compound 5

CLys = 10-6 mol/l, VLys = 1000 L, CCompound 5= 10-5 mol/l

Volume of
Compound 5 (L) / CLys
(mol/l) / CCompound 5
(mol/l) / Measured Fluorescence / RelativeEmission
Difference
0 / 0.000001 / 0 / 574.8 / 0
20 / 9.80392E-07 / 1.96E-07 / 460.3 / 114.5
40 / 9.61538E-07 / 3.85E-07 / 409.52 / 165.28
60 / 9.43396E-07 / 5.66E-07 / 358.63 / 216.17
80 / 9.25926E-07 / 7.41E-07 / 311.15 / 263.65
100 / 9.09091E-07 / 9.09E-07 / 284.29 / 290.51
150 / 8.69565E-07 / 1.3E-06 / 223.33 / 351.47
200 / 8.33333E-07 / 1.67E-06 / 182.54 / 392.26
300 / 7.69231E-07 / 2.31E-06 / 128.38 / 446.42
400 / 7.14286E-07 / 2.86E-06 / 109.25 / 465.55
500 / 6.66667E-07 / 3.33E-06 / 89.37 / 485.43
750 / 5.71429E-07 / 4.29E-06 / 77.93 / 496.87
1000 / 0.0000005 / 0.000005 / 70.63 / 504.17
Ka = 2967764 M-1,R2 = 0.9922

Table S2. Intrinsic fluorescence of BLG upon interaction with compound 5

CBLG = 10-6 mol/l, VBLG = 1000 L, CCompound 5= 10-5 mol/l

Volume of
Compound 5 (L) / CBLG
(mol/l) / CCompound 5
(mol/l) / Measured Fluorescence / RelativeEmission
Difference
0 / 0.000001 / 0 / 618.95 / 0
20 / 9.80392E-07 / 1.96E-07 / 607.11 / 11.84
40 / 9.61538E-07 / 3.85E-07 / 578.76 / 40.19
60 / 9.43396E-07 / 5.66E-07 / 555.09 / 63.86
80 / 9.25926E-07 / 7.41E-07 / 541.08 / 77.87
100 / 9.09091E-07 / 9.09E-07 / 524.27 / 94.68
150 / 8.69565E-07 / 1.3E-06 / 487.08 / 131.87
200 / 8.33333E-07 / 1.67E-06 / 448.16 / 170.79
300 / 7.69231E-07 / 2.31E-06 / 394.27 / 224.68
400 / 7.14286E-07 / 2.86E-06 / 344.2 / 274.75
500 / 6.66667E-07 / 3.33E-06 / 320.18 / 298.77
750 / 5.71429E-07 / 4.29E-06 / 256.75 / 362.2
1000 / 0.0000005 / 0.000005 / 221.84 / 397.11
Ka = 114874M-1,R2 = 0.9988

Table S3. Intrinsic fluorescence of BSA upon interaction with compound 5

CBSA = 10-6 mol/l, VBSA = 1000 L, CCompound 5= 10-5 mol/l

Volume of
Compound 5 (L) / CBSA
(mol/l) / CCompound 5
(mol/l) / Measured Fluorescence / RelativeEmission
Difference
0 / 0.000001 / 0 / 460.17 / 0
20 / 9.80392E-07 / 1.96E-07 / 434.95 / 25.22
40 / 9.61538E-07 / 3.85E-07 / 428.08 / 32.09
60 / 9.43396E-07 / 5.66E-07 / 420.33 / 39.84
80 / 9.25926E-07 / 7.41E-07 / 412.24 / 47.93
100 / 9.09091E-07 / 9.09E-07 / 397.05 / 63.12
150 / 8.69565E-07 / 1.3E-06 / 370.19 / 89.98
200 / 8.33333E-07 / 1.67E-06 / 347.43 / 112.74
300 / 7.69231E-07 / 2.31E-06 / 306.52 / 153.65
400 / 7.14286E-07 / 2.86E-06 / 279.22 / 180.95
500 / 6.66667E-07 / 3.33E-06 / 255.43 / 204.74
750 / 5.71429E-07 / 4.29E-06 / 208.01 / 252.16
1000 / 0.0000005 / 0.000005 / 179.28 / 280.89
Ka = 83471 M-1,R2 = 0.9983

Table S4. Intrinsic fluorescence of HSA upon interaction with compound 5

CHSA = 10-6 mol/l, VHSA = 1000 L, CCompound 5= 10-5 mol/l

Volume of
Compound 5 (L) / CHSA
(mol/l) / CCompound 5
(mol/l) / Measured Fluorescence / RelativeEmission
Difference
0 / 0.000001 / 0 / 526.07 / 0.0000
20 / 9.80392E-07 / 1.96E-07 / 503.02 / 23.0500
40 / 9.61538E-07 / 3.85E-07 / 496.7 / 29.3700
60 / 9.43396E-07 / 5.66E-07 / 489.89 / 36.1800
80 / 9.25926E-07 / 7.41E-07 / 481.29 / 44.7800
100 / 9.09091E-07 / 9.09E-07 / 471.03 / 55.0400
150 / 8.69565E-07 / 1.3E-06 / 452.15 / 73.9200
200 / 8.33333E-07 / 1.67E-06 / 427.39 / 98.6800
300 / 7.69231E-07 / 2.31E-06 / 385.84 / 140.2300
400 / 7.14286E-07 / 2.86E-06 / 358.44 / 167.6300
500 / 6.66667E-07 / 3.33E-06 / 334.55 / 191.5200
750 / 5.71429E-07 / 4.29E-06 / 297.12 / 228.9500
1000 / 0.0000005 / 0.000005 / 273.36 / 252.7100
Ka = 78439M-1,R2 = 0.9968

1