Supporting information for
Two cationic porphyrin isomers showing different
multimeric G-quadruplex recognition specificity against
monomeric G-quadruplexes
Xiao-Xi Huang1,2, Li-Na Zhu2,3, Bin Wu3, Yan-Fang Huo3, Na-Na Duan1,2 and De-Ming Kong1,2,*
1
State Key Laboratory of Medicinal Chemical Biology, Nankai University, Tianjin, 300071, P R
China
2
Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin
300071, P R China
3 Department
of Chemistry, Tianjin University, Tianjin, 300072, P R China
* Corresponding authors: De-Ming Kong. Tel: (+)86-22-23500938; Fax: (+)86-22-23502458;
Email: [email protected]
1. Synthesis
and
Characterization
of
5,10,15,20-tetra-{3-[2-(1-methyl-1-
piperidinyl) ethoxy] phenyl} porphyrin tetraiodide (m-TMPipEOPP)
1.1 Synthesis and characterization of 5,10,15,20-tetrakis (3-hydroxyphenyl)
porphyrin (THPP).
A suspension of 3-Hydroxybenzaldehyde (6.1 g, 0.05 mol) was dissolved in 100
mL propionic acid/DMSO (dimethyl sulfoxide) (v:v = 47:3) and the mixture was
stirred at 128 oC. Then, 3.5 mL (0.05 mol) of freshly prepared pyrrole was added. The
mixture was heated to 141 oC and refluxed for 2h, then, cooled to room temperature
and filtrated under reduced pressure. The crude residue was redissolved in CH2Cl2 or
CH2Cl2/CH3OH mixture, and purified using silica gel columns (100–200 mesh).
CH2Cl3/MeOH mixture (v:v = 7:1) was used to elute the pure product. The purple
solid of the product was obtained in 7.04% yield (0.5978g, 0.88 mmol). 1H NMR (300
MHz, [D6]DMSO, 25 ºC, TMS) (Figure S1): δ = 9.89 (s, 4H; phenolic hydroxyl H),
8.86 (s, 8H; β-pyrrole H), 7.53 (s, 12 H; Ph-H), 7.23 (s, 2H; Ph-H), 7.20 (s, 2H; Ph-H),
-2.99 ppm (s, 2H; pyrrole H); ESI:m/z: calcd for C44H30N4O4[M+H+]: 679.23; found:
679.64[M+H+]. FT-MS: m/z: calcd for C44H30N4O4: 679.23398; found: 679.2338[M].
Figure S1. 1H-NMR of 5,10,15,20-tetrakis(3-hydroxyphenyl)porphyrin (THPP). The NMR
spectra were recorded on Mercury Vx-300 spectrometer operating for 1H NMR. Chemical shifts in
the 1H NMR spectra are reported in ppm relative to the residual hydrogen atoms in the deuterated
solvents: d = 2.50 and 7.25 ppm for [D6]DMSO and CDCl3, respectively.
1.2 Synthesis and characterization of 5,10,15,20-tetra-{3-[2-(1-piperidinyl)
ethoxy]phenyl} porphyrin (TPipEOPP).
A suspension of 5,10,15,20-tetrakis (3-hydroxyphenyl) porphyrin (THPP)
(0.2172g, 0.32 mmol), 1-(2-chloroethyl)-piperidine hydrochloride (0.4714 g, 2.56
mmol), and K2CO3 (0.6192g, 4.48 mmol) in dry DMF (50 mL) was stirred for 72 h at
room temperature under N2. Then the mixture was filtered. The red-brown precipitate
was obtained and washed with DMF (10 mL) and diethyl ether (5 mL). The residue
was dissolved in dichloromethane (100 mL) and washed with water. The organic layer
was evaporated under reduced pressure. The resulting solid was isolated by
chromatography on alumina (100-200 mesh) with ethyl acetate /methanol (v:v = 50:1).
The first fraction was collected and the solvent was evaporated. Further purification
was carried out by recrystallization from CH3OH/CH2Cl2 (v:v = 20:1). The
purple-brown crystals of the product TPipEOPP were obtained in 56.9% yield (0.124
g, 0.11 mmol). 1H NMR (300 MHz, CDCl3, 25 oC, TMS) (Figure S2): δ = 8.86 (s, 8H;
β-pyrrole H), 7.80 (s, 2H; Ph-H), 7.78 (s, 6H; Ph-H), 7.61 (s, 4H; Ph-H), 7.33 (s, 2H;
Ph-H), 7.31 (s, 2H; Ph-H), 4.30 (s, 8H; OCH2), 2.88 (s, 8H; NCH2), 2.55 (s, 16H;
piperidine H), 1.61 (s, 16H; piperidine H), 1.43 (s, 8H; piperidineH), -2.81 ppm (s, 2H;
pyrrole H); ESI:m/z: calcd for C72H81N8O4[M+H+]: 1123.47; found:1124.06[M+H+].
FT-MS: m/z: calcd for C72H81N8O4: 1123.65318; found: 1123.6530[M].
Figure S2.
1H-NMR
of
5,10,15,20-tetra- {3-[2-(1-piperidinyl)ethoxy]phenyl} porphyrin
(TPipEOPP). The NMR spectra were recorded on Mercury Vx-300 spectrometer operating for 1H
NMR. Chemical shifts in the 1H NMR spectra are reported in ppm relative to the residual
hydrogen atoms in the deuterated solvents: d = 2.50 and 7.25 ppm for [D6]DMSO and CDCl3,
respectively.
1.3
Synthesis
and
characterization
of
5,10,15,20-tetra-{3-[2-(1-methyl-1-
piperidinyl) ethoxy] phenyl} porphyrin tetraiodide (m-TMPipEOPP-4I).
To a suspension of TPipEOPP (0.0584g, 0.052 mmol) in dry CH2Cl2 (35 mL) was
added CH3I (15 mL, 0.24 mmol). The mixture was stirred under N2 and heated by
using an oil bath at 40 oC for 24 h. The solvent was evaporated and the resulting solid
was washed with CH2Cl2 and diethyl ether in turn. m-TMPipEOPP-4I was obtained as
a red-purple solid in 60.7% yield (0.0355 g, 0.030 mmol). 1H NMR (300 MHz,
[D6]DMSO, 25 ºC, TMS) (Figure S1): δ = 8.90 (s, 8H; β-pyrrole H), 7.84 (s, 8H;
Ph-H), 7.78 (s, 4H; Ph-H), 7.52 (s, 4H; Ph-H), 4.69 (s, 8H; OCH2), 3.92 (s, 8H;
NCH2), 3.41–3.33 (m, 16H; piperidine H), 3.19 (s, 12H; NCH3), 2.49 (s, 8H;
piperidine H), 1.84 (s, 16H; piperidine H), -2.95 ppm (s, 2H; pyrrole H); ESI:m/z:
calcd for [C76H93N8O4-4I]/4: 295.7; found: 296.2 [M+-4I]/4. FT-MS: m/z: calcd for
[C76H 93N8 O4-4I]/4: 295.68440; found: 295.6847[M].
Figure S3. 1H-NMR of 5,10,15,20-tetra- {3-[2-(1-methyl-1-piperidinyl)ethoxy]phenyl} porphyrin
(m-TMPipEOPP). The NMR spectra were recorded on Mercury Vx-300 spectrometer operating
for 1H NMR. Chemical shifts in the 1H NMR spectra are reported in ppm relative to the residual
hydrogen atoms in the deuterated solvents: d = 2.50 and 7.25 ppm for [D6]DMSO and CDCl3,
respectively.
2. Effects of different DNAs on the UV-vis absorption spectrum of
m-TMPipEOPP
2.5
m-TMPipEOPP
(1) Hum 57
2.0
Absorbance
(2) Hum 63
(3) Hum 69
1.5
GC
LD
1.0
(1)
(2)
(3)
0.5
ssDNA2
0.0
400
500
600
700
800
Wavelength (nm)
Figure S4. UV-vis absorption spectra of m-TMPipEOPP in absence or presence of different
DNAs. Free porphyrin (black line); G-quadruplex (red line); duplex DNA (blue line);
single-stranded DNA (green line). [porphyrin] = 2.5 μM; [multimeric quadruplex] = 10 μM;
[duplex DNA] = [single-stranded DNA] = 20 μM.
3. DNA concentration-dependent changes in the absorption spectrum of
m-TMPipEOPP
3.1 Duplex DNAs
AT
Absorbance
2.0
0 M
10 M
20 M
30 M
40 M
50 M
1.5
1.0
0.5
0.0
400
500
600
700
800
Wavelength(nm)
GC
Absorbance
2.0
0 M
10 M
20 M
30 M
40 M
50 M
1.5
1.0
0.5
0.0
400
500
600
700
800
Wavelength(nm)
LD
Absorbance
2.0
0 M
10 M
20 M
30 M
40 M
50 M
1.5
1.0
0.5
0.0
400
500
600
700
800
Wavelength(nm)
Figure S5. DNA concentration-dependent absorption spectrum changes of m-TMPipEOPP in the
presence of individual duplex DNAs. The concentrations of the DNAs are labelled in the figures.
3.2 Single-stranded DNA
ssDNA2
Absorbance
2.0
0 M
10 M
20 M
30 M
40 M
50 M
1.5
1.0
0.5
0.0
400
500
600
700
800
Wavelength(nm)
Figure S6. DNA concentration-dependent absorption spectrum changes of m-TMPipEOPP in the
presence of single-stranded ssDNA2. The concentrations of ssDNA2 are labelled in the figure.
3.3 Multimeric G-quadruplexes
Hum51
2.0
2.0
1.5
1.0
Absorbance
1.5
0.5
0.0
350
1.0
400
450
500
700
750
0.10
0.05
0.5
0.00
600
650
0.0
400
500
600
700
800
Wavelength (nm)
Hum57
2.0
2.0
1.5
1.0
1.5
0.5
Absorbance
0.0
350
1.0
400
450
500
650
700
750
0.10
0.05
0.00
0.5
600
0.0
400
500
600
700
800
Wavelength (nm)
Hum63
2.0
2 .0
1 .5
1 .0
Absorbance
1.5
0 .5
0 .0
350
1.0
400
450
500
650
700
750
0.10
0.05
0.5
0.00
600
0.0
400
500
600
700
800
Wavelength (nm)
Hum69
2.0
2.0
1.5
1.0
Absorbance
1.5
0.5
0.0
350
1.0
400
450
500
650
700
750
0.10
0.05
0.00
0.5
600
0.0
400
500
600
700
800
Wavelength (nm)
Figure S7. DNA concentration-dependent absorption spectrum changes of m-TMPipEOPP in the
presence of individual multimeric G-quadurplexes. The concentrations of the G-quadruplexes are
(arrow direction): 0, 2.5, 5, 10, 15 and 20 μM.
3.4 Monomeric G-quadruplexes
2.5
M3Q
2.5
2.0
Absorbance
Oxy28
Oxy28
2.0
1.5
1.5
1.0
1.0
M
00 M
10
M
10 M
20 M
20 M
30 M
30
40M
M
40

M
50 M
50 M
0.5
0.0
0.5
400
500
600
700
800
0.0
400
500
600
700
800
Wavelength(nm)
2.5
C-MYC
2.5
Absorbance
2.0
0 M
M
10
M
10 M
20 M
M
20
30 M
M
30
40

M
40 M
50 M
50 M
KRAS
KRAS
2.0
1.5
1.5
1.0
1.0
0.5
0.0
0.5
400
500
600
700
800
0.0
400
500
600
700
800
Wavelength(nm)
Figure S8. DNA concentration-dependent absorption spectrum changes of m-TMPipEOPP in the
presence of individual monomeric G-quadurplexes (M3Q, Oxy28, C-MYC and KRAS). The
concentrations of the G-quadruplexes are labelled in the figures.
3.5 Mutants of multimeirc G-quadruplexes
Hum51-M1
2.0
Hum51-M2
2.0
2 .0
2.0
1.5
1 .5
1 .0
1.5
400
450
Absorbance
Absorbance
0 .0
350
1.0
500
0 .1 0
0 .0 5
0.5
1.0
1.5
0 .5
0.5
0.0
350
1.0
450
500
650
700
750
0.05
0.5
0 .0 0
600
650
700
0.00
750
600
0.0
0.0
400
500
600
700
800
400
Wavelength (nm)
500
800
2 .0
1 .5
1 .5
1 .0
1 .0
1.5
0 .0
350
400
450
Absorbance
0 .5
1.0
700
Hum57-M2
2.0
2 .0
1.5
600
Wavelength (nm)
Hum57-M1
2.0
Absorbance
400
0.10
500
0 .1 0
0 .0 5
0.5
0 .5
0 .0
350
1.0
650
700
500
650
700
750
0 .0 0
600
750
0.0
450
0 .0 5
0.5
0 .0 0
600
400
0 .1 0
0.0
400
500
600
700
800
400
Wavelength (nm)
600
700
800
Wavelength (nm)
Hum57-M3
2.0
500
Hum63-M1
2.0
2 .0
2 .0
1 .5
1 .5
0 .5
0 .0
350
400
450
Absorbance
Absorbance
1.5
1 .0
1.5
1 .0
500
1.0
0 .1 0
0 .0 5
0 .5
0 .0
350
450
500
0.5
0.5
0 .0 0
600
650
700
750
0.0
0.0
400
500
600
700
400
800
Hum63-M2
2.0
500
600
2 .0
1 .5
1 .5
1 .0
400
450
500
0 .0 5
Absorbance
1.0
1 .0
1.5
0 .5
0 .0
350
0 .1 0
0 .5
0 .0
350
400
450
500
650
700
750
0 .1 0
1.0
0 .0 5
0.5
0.5
0 .0 0
0 .0 0
600
800
Hum63-M3
2.0
2 .0
1.5
700
Wavelength (nm)
Wavelength (nm)
Absorbance
400
1.0
600
650
700
750
0.0
0.0
400
500
600
700
800
Wavelength (nm)
400
500
600
700
800
Wavelength (nm)
Figure S9. DNA concentration-dependent absorption spectrum changes of m-TMPipEOPP in the
presence of the mutants of multimeric G-quadurplexes. The concentrations of the G-quadruplexes
are (arrow direction): 0, 2.5, 5, 10, 15 and 20 μM.
4. DNA concentration-dependent fluorescence spectral changes of the two
porphyrin isomers in the presence of multimeric G-quadruplexes or their
mutants
600
m-TMPipEOPP + Hum51
p-TMPipEOPP + Hum51
250
Fluorescence
Fluorescence
200
25 M
400
0 M
200
25 M
150
0 M
100
50
0
600
650
700
750
0
600
800
650
Wavelength(nm)
m-TMPipEOPP + Hum57
25 M
300
0 M
150
0
600
750
800
850
p-TMPipEOPP + Hum57
200
450
Fluorescence
250
Fluorescence
600
700
Wavelength(nm)
25 M
150
0 M
100
50
650
700
750
0
600
800
650
700
750
800
850
Wavelength(nm)
Wavelength(nm)
m-TMPipEOPP + Hum63
200
p-TMPipEOPP + Hum63
450
25 M
300
0 M
Fluorescence
Fluorescence
600
0 M
100
50
150
0
600
25 M
150
650
700
750
0
600
800
650
700
750
800
850
Wavelength(nm)
Wavelength(nm)
m-TMPipEOPP + Hum69
200
p-TMPipEOPP + Hum69
600
0 M
300
150
0
600
650
700
Wavelength(nm)
750
25 M
150
Fluorescence
Fluorescence
25 M
450
800
0 M
100
50
0
600
650
700
750
Wavelength(nm)
800
850
p-TMPipEOPP + Hum51-M1
m-TMPipEOPP + Hum51-M1
200
600
25 M
400
Fluorescence
Fluorescence
25 M
0 M
200
0
600
150
0 M
100
50
650
700
750
0
600
800
650
700
750
800
850
Wavelength(nm)
Wavelength(nm)
m-TMPipEOPP + Hum51-M2
250
p-TMPipEOPP + Hum51-M2
450
200
Fluorescence
Fluorescence
25 M
300
0 M
150
25 M
150
0 M
100
50
0
600
650
700
750
0
600
800
650
m-TMPipEOPP + Hum57-M1
250
0 M
300
150
0
600
650
700
750
25 M
150
0 M
100
0
600
800
650
700
750
800
850
Wavelength(nm)
m-TMPipEOPP + Hum57-M2
25 M
300
0 M
250
p-TMPipEOPP + Hum57-M2
25 M
200
Fluorescence
Fluorescence
850
50
450
150
0
600
800
p-TMPipEOPP + Hum57-M1
Wavelength(nm)
600
750
200
25 M
450
Fluorescence
Fluorescence
600
700
Wavelength(nm)
wavelength(nm)
Wavelength(nm)
150
0 M
100
50
650
700
750
Wavelength(nm)
800
0
600
650
700
750
Wavelength(nm)
800
850
m-TMPipEOPP + Hum57-M3
300
250
450
25 M
25 M
300
Fluorescence
Fluorescence
p-TMPipEOPP + Hum57-M3
0 M
150
200
0 M
150
100
50
0
600
600
650
700
750
0
600
800
700
750
800
Wavelength(nm)
m-TMPipEOPP + Hum63-M1
p-TMPipEOPP + Hum63-M1
850
200
Fluorescence
25 M
Fluorescence
650
Wavelength(nm)
400
0 M
200
25 M
150
0 M
100
50
0
600
650
700
750
0
600
800
650
600
700
750
m-TMPipEOPP + Hum63-M2
250
300
0 M
Fluorescence
Fluorescence
25 M
150
650
700
750
800
150
0 M
100
450
25 M
300
0 M
150
650
700
Wavelength(nm)
0
600
650
700
750
Wavelength(nm)
m-TMPipEOPP + Hum63-M3
Fluorescence
25 M
50
Wavelength(nm)
0
600
850
p-TMPipEOPP + Hum63-M2
200
450
0
600
800
Wavelength(nm)
Wavelength(nm)
750
800
800
850
p-TMPipEOPP + Hum63-M3
Fluorescence
250
200
25 M
150
0 M
100
50
0
600
650
700
750
800
850
Wavelength(nm)
Figure S10. DNA concentration-dependent fluorescence spectrum changes of m-TMPipEOPP
(Left) and p-TMPipEOPP (Right) in the presence of multimeric G-quadurplexes and their
mutants. When m-TMPipEOPP was used, the concentrations of the G-quadruplexes are (arrow
direction): 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20 and 25 μM. When p-TMPipEOPP was used, the
concentrations of the G-quadruplexes are (arrow direction): 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8,
0.9, 1, 1.5, 2, 2.5, 3, 3.5, 4, 5, 6, 7, 8, 9, 10, 15, 20 and 25 μM.
200
160
F674nm-F709nm
120
Hum45
80
40
0
Slope(Hum45)/Slope(Hum21)=891
-40
-80
Hum21
-120
0
10
20
30
40
50
G-rich sequence concentration (M)
Figure S11. Comparison of fluorescence signal changes (F674nm-F709nm) in the presence of
multimeric G-quadruplex Hum45 and monomeric G-quadruplex Hum21.
5. Job Plot analysis for the interactions between p-TMPipEOPP and
G-quadruplexes
5.1 Monomeric G-quadruplex Hum21
Hum21
(a)
(b)
Hum21
0.4
1.6
0.3
A454
A421
1.2
0.8
0.4
0.2
0.1
0.32
0.33
0.0
0.0
0.0
0.2
0.4
0.6
0.8
1.0
0.0
(c)
Hum21
0.2
0.4
0.6
0.8
1.0
[o-TMPipEOPP]/([o-TMPipEOPP]+[Hum21])
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum21])
(d)
Hum21
400
0.3
Fluorescence
A695
300
0.2
0.1
200
100
0.34
0.33
0.0
0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum21])
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum21])
Figure S12. Job plot analysis of the interaction between p-TMPipEOPP and Hum21 utilizing the
absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at 719
nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum21] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum21] = 5 μM, (c) [p-TMPipEOPP] + [Hum21] = 30 μM.
5.2 Multimeric G-quadruplex Hum45
(a)
Hum45
(b)
1.6
Hum45
0.9
1.2
A454
A421
0.6
0.8
0.3
0.4
0.39
0.37
0.0
0.0
0.0
0.2
0.4
0.6
0.8
1.0
0.0
([p-TMPipEOPP]/(p-TMPipEOPP]+[Hum45]))
(c)
Hum45
(d)
0.4
Fluorescence
A695
0.3
0.2
0.1
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/(p-TMPipEOPP]+[Hum45])
Hum45
800
600
400
200
0.38
0.38
0.0
0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/(p-TMPipEOPP]+[Hum45])
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/(p-TMPipEOPP]+[Hum45])
Figure S13. Job plot analysis of the interaction between p-TMPipEOPP and Hum45 utilizing the
absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at 719
nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum45] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum45] = 5 μM, (c) [p-TMPipEOPP] + [Hum45] = 30 μM.
5.3 Multimeric G-quadruplex Hum51
(a)
Hum51
1.6
A421
1.2
0.8
0.4
0.42
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51])
Hum51
(b)
0.6
A454
0.4
0.2
0.43
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51])
(c)
Hum51
0.4
A695
0.3
0.2
0.1
0.41
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51])
(d)
Hum51
Fluorescence
800
600
400
200
0.41
0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51])
Figure S14. Job plot analysis of the interaction between p-TMPipEOPP and Hum51 utilizing the
absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at 719
nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum51] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum51] = 5 μM, (c) [p-TMPipEOPP] + [Hum51] = 30 μM.
5.4 Multimeric G-quadruplex Hum57
(a)
Hum57
1.6
(b)
Hum57
0.6
0.4
A454
A421
1.2
0.8
0.2
0.4
0.45
0.46
0.0
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57])
(c)
0.0
0.2
0.4
0.6
0.8
1.0
[o-TMPipEOPP]/([o-TMPipEOPP]+[Hum57])
Hum57
0.4
A695
0.3
0.2
0.1
0.46
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57])
(d)
Hum57
Fluorescence
800
600
400
200
0.45
0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57])
Figure S15. Job plot analysis of the interaction between p-TMPipEOPP and Hum57 utilizing the
absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at 719
nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum57] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum57] = 5 μM, (c) [p-TMPipEOPP] + [Hum57] = 30 μM.
5.5 Multimeric G-quadruplex Hum63
(a)
Hum63
1.6
A421
1.2
0.8
0.4
0.50
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63])
(c)
Hum63
Hum63
0.8
0.4
0.6
0.3
A695
A454
(b)
0.4
0.2
0.2
0.1
0.52
0.51
0.0
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63])
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63])
Hum63
(d)
Fluorescence
1200
900
600
0.50
300
0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63])
Figure S16. Job plot analysis of the interaction between p-TMPipEOPP and Hum63 utilizing the
absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at 719
nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum63] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum63] = 5 μM, (c) [p-TMPipEOPP] + [Hum63] = 30 μM.
5.6 Multimeric G-quadruplex Hum69
Hum69
(a)
A421
1.2
0.8
0.4
0.49
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum69])
(b)
Hum69
0.8
A454
0.6
0.4
0.2
0.49
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[o-TMPipEOPP]/([o-TMPipEOPP]+[Hum72])
(c)
Hum69
(d)
0.5
Hum69
900
Fluorescence
A695
0.4
0.3
0.2
600
300
0.1
0.50
0.48
0.0
0
0.0
0.2
0.4
0.6
0.8
1.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum69])
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum69])
Figure S17. Job plot analysis of the interaction between p-TMPipEOPP and Hum69 utilizing the
absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at 719
nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum69] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum69] = 5 μM, (c) [p-TMPipEOPP] + [Hum69] = 30 μM.
5.7 Multimeric G-quadruplex Hum51-M1
(a)
Hum51-M1
(b)
1.6
0.3
A454
1.2
A421
Hum51-M1
0.4
0.8
0.4
0.2
0.1
0.38
0.0
0.0
0.2
0.4
0.6
0.37
0.0
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51-M1])
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51-M1])
(c)
Hum51-M1
0.4
A695
0.3
0.2
0.1
0.38
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51-M1])
Fluorescence
(d)
Hum51-M1
600
400
200
0.38
0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51-M1])
Figure S18. Job plot analysis of the interaction between p-TMPipEOPP and Hum51-M1 utilizing
the absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at
719 nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum51-M1] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum51-M1] = 5 μM, (c) [p-TMPipEOPP] + [Hum51-M1] = 30 μM.
5.8 Multimeric G-quadruplex Hum57-M1
(a)
Hum57-M1
Hum57-M1
(b)
1.6
0.6
0.4
A454
A421
1.2
0.8
0.2
0.4
0.40
0.38
0.0
0.0
0.2
0.4
0.6
0.0
0.8
1.0
0.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M1])
(c)
Hum57-M1
0.4
0.6
0.8
1.0
Hum57-M1
(d)
0.4
Fluorescence
900
0.3
A695
0.2
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M1])
0.2
0.1
600
300
0.37
0.39
0
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M1])
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M1])
Figure S19. Job plot analysis of the interaction between p-TMPipEOPP and Hum57-M1 utilizing
the absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at
719 nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum57-M1] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum57-M1] = 5 μM, (c) [p-TMPipEOPP] + [Hum57-M1] = 30 μM.
5.9 Multimeric G-quadruplex Hum63-M1
(a)
Hum63-M1
0.6
A454
A421
1.2
0.8
0.4
0.2
0.4
0.39
0.38
0.0
0.0
0.2
0.4
0.6
0.0
0.8
0.0
1.0
(c)
0.2
(d)
Hum63-M1
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M1])
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M1])
Hum63-M1
1200
0.4
Fluorescence
0.3
A695
Hum63-M1
(b)
1.6
0.2
0.1
900
600
300
0.39
0.4
0.0
0
0.0
0.2
0.4
0.6
0.8
0.0
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M1])
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M1])
Figure S20. Job plot analysis of the interaction between p-TMPipEOPP and Hum63-M1 utilizing
the absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at
719 nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum63-M1] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum63-M1] = 5 μM, (c) [p-TMPipEOPP] + [Hum63-M1] = 30 μM.
5.10
Multimeric G-quadruplex Hum51-M2
(a)
(b)
Hum51-M2
1.6
Hum51-M2
0.6
1.2
A454
A421
0.4
0.8
0.2
0.4
0.46
0.45
0.0
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51-M2])
(c)
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51-M2])
Hum51-M2
0.4
A695
0.3
0.2
0.1
0.46
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51-M2])
Fluorescence
(d)
Hum51-M2
600
400
200
0.45
0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum51-M2])
Figure S21. Job plot analysis of the interaction between p-TMPipEOPP and Hum51-M2 utilizing
the absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at
719 nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum51-M2] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum51-M2] = 5 μM, (c) [p-TMPipEOPP] + [Hum51-M2] = 30 μM.
5.11
(a)
Multimeric G-quadruplex Hum57-M2
Hum57-M2
Hum57-M2
(b)
1.6
0.6
A454
A421
1.2
0.8
0.4
0.2
0.4
0.44
0.44
0.0
0.0
0.0
0.2
0.4
0.6
0.8
0.0
1.0
(c)
(d)
Hum57-M2
0.4
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M2])
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M2])
Hum57-M2
900
Fluorescence
A695
0.3
0.2
0.1
600
300
0.44
0.44
0
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M2])
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M2])
Figure S22. Job plot analysis of the interaction between p-TMPipEOPP and Hum57-M2 utilizing
the absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at
719 nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum57-M2] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum57-M2] = 5 μM, (c) [p-TMPipEOPP] + [Hum57-M2] = 30 μM.
5.12
(a)
Multimeric G-quadruplex Hum63-M2
Hum63-M2
0.6
A454
1.2
A421
Hum63-M2
(b)
1.6
0.8
0.4
0.2
0.4
0.46
0.47
0.0
0.0
0.0
0.2
0.4
0.6
0.8
0.0
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M2])
Hum63-M2
(c)
(d)
0.4
0.4
0.6
0.8
1.0
Hum63-M2
1200
Fluorescence
900
0.3
A695
0.2
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M2])
0.2
0.1
600
300
0.47
0.46
0.0
0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M2])
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M2])
Figure S23. Job plot analysis of the interaction between p-TMPipEOPP and Hum63-M2 utilizing
the absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at
719 nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum63-M2] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum63-M2] = 5 μM, (c) [p-TMPipEOPP] + [Hum63-M2] = 30 μM.
5.13
Multimeric G-quadruplex Hum57-M3
Hum57-M3
(a)
(b)
Hum57-M3
0.8
1.2
A421
0.6
A454
0.8
0.4
0.4
0.2
0.5
0.0
0.0
0.2
0.4
0.6
0.5
0.0
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M3])
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M3])
Hum57-M3
(c)
0.4
A695
0.3
0.2
0.1
0.51
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M3])
(d)
Hum57-M3
Fluorescence
1200
800
400
0.5
0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M3])
Figure S24. Job plot analysis of the interaction between p-TMPipEOPP and Hum57-M3 utilizing
the absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at
719 nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum57-M3] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum57-M3] = 5 μM, (c) [p-TMPipEOPP] + [Hum57-M3] = 30 μM.
5.14
(a)
Multimeric G-quadruplex Hum63-M3
Hum63-M3
(b)
1.6
0.8
0.6
A454
A421
1.2
Hum63-M3
0.8
0.4
0.4
0.2
0.50
0.51
0.0
0.0
0.2
0.4
0.6
0.8
0.0
0.0
1.0
(c)
Hum63-M3
0.5
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M3])
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M3])
(d)
Hum63-M3
1200
Fluorescence
0.4
A695
0.3
0.2
900
600
300
0.1
0.51
0.5
0.0
0
0.0
0.2
0.4
0.6
0.8
1.0
0.0
0.2
0.4
0.6
0.8
1.0
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M3])
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum63-M3])
[p-TMPipEOPP]/([p-TMPipEOPP]+[Hum57-M3])
Figure S25. Job plot analysis of the interaction between p-TMPipEOPP and Hum63-M3 utilizing
the absorption signals at (a) 421 nm, (b) 454 nm, (c) 695 nm and (d) the fluorescence signals at
719 nm (λex = 700 nm), respectively. (a) [p-TMPipEOPP] + [Hum63-M3] = 2 μM, (b) and (d)
[p-TMPipEOPP] + [Hum63-M3] = 5 μM, (c) [p-TMPipEOPP] + [Hum63-M3] = 30 μM.
6. Job Plot analysis for the interactions between m-TMPipEOPP and
G-quadruplexes
6.1 Monomeric G-quadruplex Hum21
Hum21
0.16
0.14
0.12
A452
0.10
0.08
0.06
0.04
0.02
0.00
-0.02
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum21])
Hum21
0.08
A666
0.06
0.04
0.02
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum21])
Figure S26. Job plot analysis of the interaction between m-TMPipEOPP and Hum21 utilizing the
absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum21] = 10 μM.
6.2 Multimeric G-quadruplex Hum45
Hum 45
0.25
A452
0.20
0.15
0.10
0.05
0.31
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum45])
Hum45
0.06
0.05
A666
0.04
0.03
0.02
0.01
0.31
0.00
-0.01
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum45])
Figure S27. Job plot analysis of the interaction between m-TMPipEOPP and Hum45 utilizing the
absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum45] = 10 μM.
6.3 Multimeric G-quadruplex Hum51
Hum 51
0.4
A452
0.3
0.2
0.1
0.33
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum51])
Hum51
0.06
A666
0.04
0.02
0.33
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum51])
Figure S28. Job plot analysis of the interaction between m-TMPipEOPP and Hum51 utilizing the
absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum51] = 10 μM.
6.4 Multimeric G-quadruplex Hum57
Hum 57
A452
0.6
0.4
0.2
0.33
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum57])
Hum57
0.12
0.10
A666
0.08
0.06
0.04
0.02
0.33
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum57])
Figure S29. Job plot analysis of the interaction between m-TMPipEOPP and Hum57 utilizing the
absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum57] = 10 μM.
6.5 Multimeric G-quadruplex Hum63
Hum 63
0.5
A452
0.4
0.3
0.2
0.1
0.33
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum63])
Hum63
0.06
0.05
A666
0.04
0.03
0.02
0.01
0.33
0.00
-0.01
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum63])
Figure S30. Job plot analysis of the interaction between m-TMPipEOPP and Hum63 utilizing the
absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum63] = 10 μM.
6.6 Multimeric G-quadruplex Hum69
Hum69
0.3
A452
0.2
0.1
0.33
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum69])
Hum69
0.08
A666
0.06
0.04
0.02
0.33
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum69])
Figure S31. Job plot analysis of the interaction between m-TMPipEOPP and Hum69 utilizing the
absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum69] = 10 μM.
6.7 Multimeric G-quadruplex Hum51-M1
Hum51-M1
0.40
0.35
0.30
A452
0.25
0.20
0.15
0.10
0.05
0.29
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum51-M1])
Hum51-M1
0.06
0.05
A666
0.04
0.03
0.02
0.01
0.31
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum51-M1])
Figure S32. Job plot analysis of the interaction between m-TMPipEOPP and Hum51-M1 utilizing
the absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum51-M1] = 10
μM.
6.8 Multimeric G-quadruplex Hum57-M1
Hum57-M1
0.45
0.40
0.35
A452
0.30
0.25
0.20
0.15
0.10
0.31
0.05
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum57-M1])
Hum57-M1
0.10
A666
0.08
0.06
0.04
0.31
0.02
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum57-M1])
Figure S33. Job plot analysis of the interaction between m-TMPipEOPP and Hum57-M1 utilizing
the absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum57-M1] = 10
μM.
6.9 Multimeric G-quadruplex Hum63-M1
Hum63-M1
0.40
0.35
0.30
A452
0.25
0.20
0.15
0.10
0.31
0.05
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum63-M1])
Hum63-M1
0.08
0.07
0.06
A666
0.05
0.04
0.03
0.02
0.01
0.31
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum63-M1])
Figure S34. Job plot analysis of the interaction between m-TMPipEOPP and Hum63-M1 utilizing
the absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum63-M1] = 10
μM.
Multimeric G-quadruplex Hum51-M2
Hum51-M2
0.4
0.3
A452
6.10
0.2
0.1
0.33
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum51-M2])
Hum51-M2
0.08
A666
0.06
0.04
0.02
0.33
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum51-M2])
Figure S35. Job plot analysis of the interaction between m-TMPipEOPP and Hum51-M2 utilizing
the absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum51-M2] = 10
μM.
Multimeric G-quadruplex Hum57-M2
Hum57-M2
0.5
0.4
0.3
A452
6.11
0.2
0.1
0.0
0.33
-0.1
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum57-M2])
Hum57-M2
0.12
0.10
A666
0.08
0.06
0.04
0.02
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum57-M2])
Figure S36. Job plot analysis of the interaction between m-TMPipEOPP and Hum57-M2 utilizing
the absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum57-M2] = 10
μM.
Multimeric G-quadruplex Hum63-M2
Hum63-M2
0.3
A452
6.12
0.2
0.1
0.33
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum63-M2])
Hum63-M2
0.10
0.08
A666
0.06
0.04
0.02
0.33
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum63-M2])
Figure S37. Job plot analysis of the interaction between m-TMPipEOPP and Hum63-M2 utilizing
the absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum63-M2] = 10
μM.
Multimeric G-quadruplex Hum57-M3
Hum57-M3
0.5
0.4
0.3
A452
6.13
0.2
0.1
0.33
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum57-M3])
Hum57-M3
0.12
0.10
A666
0.08
0.06
0.04
0.33
0.02
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum57-M3])
Figure S38. Job plot analysis of the interaction between m-TMPipEOPP and Hum57-M3 utilizing
the absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum57-M3] = 10
μM.
Multimeric G-quadruplex Hum63-M3
Hum63-M3
0.5
0.4
0.3
A452
6.14
0.2
0.1
0.33
0.0
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum63-M3])
Hum63-M3
0.06
0.05
A666
0.04
0.03
0.02
0.01
0.32
0.00
0.0
0.2
0.4
0.6
0.8
1.0
[m-TMPipEOPP]/([m-TMPipEOPP]+[Hum63-M3])
Figure S39. Job plot analysis of the interaction between m-TMPipEOPP and Hum63-M3 utilizing
the absorption signals at 452 nm and 666 nm, respectively. [m-TMPipEOPP] + [Hum63-M3] = 10
μM.
7. Circular dichroism (CD) spectroscopy of monomeric and multimeric
G-quadruplexes
7.1 Under dilute conditions
4
Hum 69
Hum 63
Hum 57
Hum 51
Hum 45
Hum 21
4
3
CD/mdeg
CD/mdeg
6
2
Hum 45
Hum 51
Hum 51-M1
Hum 51-M2
2
1
0
0
-1
-2
220
240
260
280
Wavelength(nm)
300
320
220
240
260
280
Wavelength(nm)
300
320
6
6
CD/mdeg
CD/mdeg
4
8
Hum 59
Hum 59-M1
Hum 59-M2
Hum 59-M3
2
0
220
4
Hum63
Hum63-M1
Hum63-M2
Hum63-M3
Hum69
2
0
240
260
280
300
-2
220
320
240
260
280
300
320
Wavelength(nm)
Wavelength(nm)
Figure S40. CD spectra of monomeric and multimeric G-quadruplexes under dilute conditions
7.2 Under molecular crowding conditions
20
Hum69
Hum63
Hum57
Hum51
Hum45
Hum21
10
8
CD/mdeg
CD/mdeg
15
5
4
0
0
-5
-4
220
240
260
280
Wavelength (nm)
300
Hum45
Hum51
Hum51-M1
Hum51-M2
12
320
220
240
260
280
Wavelength(nm)
300
320
16
25
CD/mdeg
12
8
4
15
10
5
0
0
-5
-4
220
Hum63
Hum63-M1
Hum63-M2
Hum63-M3
Hum69
20
CD/mdeg
Hum57
Hum57-M1
Hum57-M2
Hum57-M3
240
260
280
300
Hum 45
Hum 51
Hum 51-M1
Hum 51-M2
12
8
CD/mdeg
Wavelength(nm)
4
0
-4
220
240
260
280
300
320
Wavelength(nm)
320
-10
220
240
260
280
300
320
Wavelength (nm)
Figure S41. CD spectra of monomeric and multimeric G-quadruplexes under molecular crowding
conditions
8. Scatchard
analysis of the interactions between m-TMPipEOPP and
multimeric G-quadruplexes
Hum45
Hum51
0.32
0.20
0.30
0.19
0.28
0.26
0.18
r/Cf
r/Cf
0.21
0.24
0.17
0.22
0.16
0.20
0.15
0.18
0.28
0.29
0.30
0.31
0.32
0.16
0.33
0.32
0.34
0.36
0.40
Hum63
Hum57
0.23
0.38
r
r
0.60
0.22
0.55
0.21
r/Cf
r/Cf
0.20
0.19
0.50
0.45
0.18
0.17
0.40
0.16
0.35
0.15
0.28
0.30
0.32
0.30
0.34
0.32
0.34
0.38
0.40
Hum51-M1
Hum69
0.44
0.36
r
r
0.40
0.18
0.36
0.17
r/Cf
r/Cf
0.19
0.16
0.32
0.15
0.28
0.14
0.24
0.32
0.34
0.21
0.36
0.38
0.28
0.40
0.30
0.31
r
r
Hum51-M2
Hum57-M1
0.32
0.33
0.28
0.20
0.19
0.26
0.18
r/Cf
r/Cf
0.29
0.17
0.16
0.24
0.22
0.15
0.20
0.14
0.13
0.18
0.28
0.29
0.30
0.31
r
0.32
0.33
0.34
0.27
0.28
0.29
0.30
r
0.31
0.32
0.33
Hum57-M2
0.30
0.28
0.23
0.26
0.22
r/Cf
0.24
r/Cf
Hum57-M3
0.24
0.22
0.20
0.21
0.20
0.19
0.18
0.16
0.18
0.14
0.17
0.40
0.41
0.42
0.43
0.28
0.44
0.29
0.30
0.31
r
Hum63-M1
0.42
0.33
0.34
0.33
0.34
Hum63-M2
0.24
0.40
0.38
0.23
0.36
0.22
r/Cf
r/Cf
0.32
r
0.34
0.21
0.32
0.20
0.30
0.19
0.28
0.26
0.26
0.18
0.27
0.28
0.29
0.30
0.31
0.32
0.33
0.28
0.29
0.30
0.31
0.32
r
r
Hum63-M3
0.24
0.23
r/Cf
0.22
0.21
0.20
0.19
0.18
0.17
0.28
0.29
0.30
0.31
0.32
0.33
0.34
0.35
r
Figure S42. Scatchard plots for m-TMPipEOPP with multimeric G-quadruplexes. The change in
the absorbance difference between 452 and 418 nm as a function of G-quadruplex concentration
was used to construct Scatchard plots
Table S1. Binding parameters for the interactions between m-TMPipEOPP and multimeric
G-quadruplexes
na
Ka
(×10-6 M-1)b
na
Ka
(×10-6 M-1)b
Hum 21
undetected
undetected
Hum 51-M2
0.46
1.10
Hum45
0.46
1.11
Hum 57-M1
0.48
1.30
Hum51
0.50
1.66
Hum 57-M2
0.47
1.38
Hum57
0.48
1.05
Hum 57-M3
0.50
1.08
Hum63
0.52
2.53
Hum 63-M1
0.46
2.08
Hum69
0.52
2.05
Hum 63-M2
0.50
1.10
Hum 51-M1
0.45
1.09
Hum 63-M3
0.50
1.08
G-quadruplex
G-quadruplex
a
n is the number of m-TMPipEOPP-binding sites on the G-quadruplex
b
Ka is the binding constant for the interaction between m-TMPipEOPP and multimeric
G-quadurplexes
9. Two possible binding modes between m-TMPipEOPP and multimeric
G-quadruplexes
Scheme S1. Two possible binding modes between m-TMPipEOPP and multimeric
G-quadruplexes.
10. G-quadruplex-stabilizing abilities of m-TMPipEOPP to G-quadruplexes
10.1
Under dilute conditions
Hum 45
Hum 21
1.0
Normalized absorbance
Normalized absorbance
1.0
0.8
o
60.8 C
0.6
0.4
o
60.4 C
0.2
0.0
0.8
o
54.9 C
0.6
0.4
o
45.8 C
0.2
0.0
20
30
40
50
60
70
80
20
30
o
40
50
80
Hum 57
1.0
Normalized absorbance
Normalized absorbance
70
Temperature ( C)
Hum 51
0.8
o
55.6 C
0.6
0.4
o
47.6 C
0.2
0.0
1.0
0.8
o
59.0 C
0.6
0.4
o
49.8 C
0.2
0.0
20
30
40
50
60
70
20
30
o
40
50
60
70
80
o
Temperature ( C)
Temperature ( C)
Hum 63
Hum 69
1.0
1.0
Normalized absorbance
Normalized absorbance
60
o
Temperature ( C)
0.8
o
60.3 C
0.6
0.4
o
51.4 C
0.2
0.0
0.8
o
55.2 C
0.6
0.4
o
48.6 C
0.2
0.0
20
30
40
50
60
o
Temperature ( C)
70
80
20
30
40
50
60
70
80
o
Temperature ( C)
Figure S43. Melting temperature (T1/2) detection of G-quadruplexes in the absence (black) and
presence (red) of 5 μM m-TMPipEOPP under dilute conditions. Scatter: experimental data, line:
fitting curves.
10.2
Under molecular crowding conditions
Hum 21
Hum 45
Normalized absorbance
Normalized absorbance
1.0
0.8
o
70.6 C
0.6
0.4
o
69.6 C
0.2
0.0
1.0
0.8
o
56.6 C
0.6
0.4
o
51.5 C
0.2
0.0
20
30
40
50
60
70
80
90
20
30
40
70
80
Hum 57
Hum 51
1.0
Normalized absorbance
Normalized absorbance
60
Temperature ( C)
Temperature ( C)
0.8
o
57.9 C
0.6
0.4
o
53.1 C
0.2
1.0
0.8
o
62.4 C
0.6
0.4
o
56.0 C
0.2
0.0
0.0
20
30
40
50
60
70
20
80
30
o
40
50
60
70
80
o
Temperature ( C)
Temperature ( C)
Hum 63
Hum 69
1.0
Normalized absorbance
1.0
Normalized absorbance
50
o
o
0.8
o
63.4 C
0.6
0.4
o
57.4 C
0.2
0.8
o
58.1 C
0.6
0.4
o
52.2 C
0.2
0.0
0.0
20
30
40
50
60
70
o
Temperature ( C)
80
20
30
40
50
60
70
80
90
o
Temperature ( C)
Figure S44. Melting temperature (T1/2) detection of G-quadruplexes in the absence (black) and
presence (red) of 5 μM m-TMPipEOPP under molecular crowding conditions. Scatter:
experimental data, line: fitting curves.
11. Stabilities of multimeric G-quadruplexes with different pocket sizes
Under dilute conditions
Hum51
Hum51-M1
Hum51-M2
Normalized absorbance
1.0
0.8
o
0.6
51.3 C
0.4
o
45.8 C
o
47.6 C
0.2
0.0
20
30
40
50
60
70
o
Normalized absorbance
Temperature ( C)
Hum57
Hum57-M1
Hum57-M2
Hum57-M3
1.0
0.8
o
0.6
51.4 C
o
0.4
52.6 C
o
45.6 C
o
49.8 C
0.2
0.0
20
30
40
50
60
70
o
Temperature ( C)
Normalized absorbance
11.1
Hum63
Hum63-M1
Hum63-M2
Hum63-M3
1.0
0.8
o
0.6
51.4 C
o
0.4
52.9 C
o
45.8 C
o
50.9 C
0.2
0.0
20
30
40
50
60
o
Temperature ( C)
70
Figure S45. Melting temperature detection of multimeric G-quadruplexes with different pocket
sizes under dilute conditions. Scatter: experimental data, line: fitting curves.
Normalized absorbance
Under molecular crowding conditions
1.0
Hum51
Hum51-M1
Hum51-M2
0.8
0.6
0.4
o
o
55.2 C
51.4 C
o
0.2
53.1 C
0.0
20
30
40
50
60
70
80
o
Temperature ( C)
Hum57
Hum57-M1
Hum57-M2
Hum57-M3
Normalized absorbance
1.0
0.8
o
0.6
57.5 C
o
0.4
56.0 C
o
50.7 C
o
55.0 C
0.2
0.0
20
30
40
50
60
70
80
o
Temperature ( C)
Hum63
Hum63-M1
Hum63-M2
Hum63-M3
1.0
Normalized absorbance
11.2
0.8
o
0.6
57.4 C
o
57.6 C
o
0.4
51.5 C
o
54.8 C
0.2
0.0
20
30
40
50
60
70
o
Temperature ( C)
80
Figure S46. Melting temperature detection of multimeric G-quadruplexes with different pocket
sizes under molecular crowding conditions. Scatter: experimental data, line: fitting curves.