Acetylpyrene-labelled 7-methylguanine nucleotides: unusual ... · R. Kasprzyk et al. 2 m7GDP...
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R. Kasprzyk et al. 1 Acetylpyrene-labelled 7-methylguanine nucleotides: unusual fluorescence properties and application to Decapping Scavenger activity monitoring Renata Kasprzyk 1,2,3 , Joanna Kowalska 1 , Zbigniew Wieczorek 4 , Mariusz Szabelski 4 , Ryszard Stolarski 1 , Jacek Jemielity 2 1 Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland 2 Centre of New Technologies, University of Warsaw, Poland 3 College of Inter-Faculty Individual Studies in Mathematics and Natural Sciences, University of Warsaw, Zwirki i Wigury 93, 02-089, Warsaw, Poland 4 Department of Physics and Biophysics, University of Warmia and Mazury in Olsztyn, Oczapowskiego 4, 10-719 Olsztyn, Poland Supporting Information (SI2): HPLC profiles, HR MS, NMR and fluorescence spectra Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry. This journal is © The Royal Society of Chemistry 2016
Transcript of Acetylpyrene-labelled 7-methylguanine nucleotides: unusual ... · R. Kasprzyk et al. 2 m7GDP...
R. Kasprzyk et al.
1
Acetylpyrene-labelled 7-methylguanine nucleotides: unusual fluorescence properties and application to Decapping Scavenger activity monitoring
Renata Kasprzyk1,2,3, Joanna Kowalska1, Zbigniew Wieczorek4, Mariusz Szabelski4, Ryszard Stolarski1, Jacek Jemielity2
1 Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland 2 Centre of New Technologies, University of Warsaw, Poland 3College of Inter-Faculty Individual Studies in Mathematics and Natural Sciences, University of Warsaw, Zwirki i Wigury 93, 02-089, Warsaw, Poland 4Department of Physics and Biophysics, University of Warmia and Mazury in Olsztyn, Oczapowskiego 4, 10-719 Olsztyn, Poland
Supporting Information (SI2): HPLC profiles, HR MS, NMR and fluorescence spectra
Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry.This journal is © The Royal Society of Chemistry 2016
R. Kasprzyk et al.
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m7GDPβS-pyrene/NH4+ (1a)
Form
ula
Sp
ectr
osc
op
ic p
rop
erti
es
absorbance emission λexc = 282 nm emission λexc = 369 nm excitation at λem = 385 nm excitation at λexc = 471 nm
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HP
LC
rea
ctio
n p
rofi
le
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4
HP
LC
pro
file
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5
HR
MS
(-)
ES
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1H
NM
R (
400 M
Hz,
D2O
, 25°C
)
m7GDPbetaS_piren_NH4_1H_D2O_25deg.esp
8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0
Chemical Shift (ppm)
0
0.005
0.010
0.015
0.020
0.025
0.030
0.035
0.040
0.045
0.050
0.055
0.060
0.065
Norm
alized Inte
nsity
3.001.200.991.051.050.421.730.890.992.700.922.722.64
8.0
68.0
38.0
1
7.9
07.8
97.8
77.5
87.5
7 7.5
47.5
3
4.7
24.6
64.5
0 4.4
9 4.3
8 4.3
44.1
34.1
14.0
5 3.9
33.9
23.9
03.6
43.6
3
3.1
2
H1’
H5’’
H5’
H4’
H3’
H2’
-CH3 Water
aromatic (H(N1), H2-10)
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31P
NM
R (
162 M
Hz,
D2O
, 25°C
)
m7GDPbetaS_piren_31P_D2O_25deg
8 6 4 2 0 -2 -4 -6 -8 -10 -12
Chemical Shift (ppm)
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
Norm
alized Inte
nsity
1.001.00
7.3
77.3
07.1
27.0
5
-11.9
8-1
2.1
6
Pα
Pβ
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m7GTPγS-pyrene/NH4+ (1b)
Form
ula
Sp
ectr
osc
op
ic p
rop
erti
es
absorbance emission λexc = 282 nm emission λexc = 369 nm excitation at λem = 385 nm excitation at λexc = 471 nm
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HP
LC
rea
ctio
n p
rofi
le
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HP
LC
pro
file
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HR
MS
(-)
ES
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1H
NM
R (
50
0 M
Hz,
D2O
, 25°C
)
m7GTPSpir_D2O_6mg_500trp_1H_25C.esp
8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5
Chemical Shift (ppm)
0
0.05
0.10
0.15
Norm
alized Inte
nsity
3.000.951.001.081.231.271.151.002.030.980.903.860.970.88
8.3
78.3
6
8.1
6
8.0
68.0
57.9
47.8
2 7.6
97.6
87.6
6
4.8
44.7
54.7
3 4.4
64.4
64.4
5 4.2
0
4.1
94.1
3 3.9
9 3.9
83.8
13.7
3
3.2
4
H1’
H5’’
H5’
H4’
H3’
H2’
H3
H2 H5
H8
H10
H9
H7, H6, H4,
-CH2-
-CH3
Water
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31P
NM
R (
162 M
Hz,
D2O
, 25°C
)
m7GTPgammaS_piren_31P_D2O_25deg.esp
5 0 -5 -10 -15 -20 -25
Chemical Shift (ppm)
-0.1
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
Norm
alized Inte
nsity
1.001.060.97
7.6
67.5
1
-11.5
1-1
1.6
2
-23.1
1-2
3.2
3-2
3.3
9
Pγ
Pα
Pβ
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2D
TO
CS
Y N
MR
(500 M
Hz,
D2O
, 25°C
)
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2D
CO
SY
NM
R (
500 M
Hz,
D2O
, 25°C
)
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2D
NO
ES
Y N
MR
(500 M
Hz,
D2O
, 25°C
)
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2D
RO
ES
y N
MR
(500 M
Hz,
D2O
, 25°C
)
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gC
hsq
c (1
H-1
3C
) N
MR
(500 M
Hz,
D2O
, 25°C
)
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m7GpCH2ppS-pyrene/TEA+ (1c) F
orm
ula
Sp
ectr
osc
op
ic p
rop
erti
es
absorbance emission λexc = 282 nm emission λexc = 369 nm excitation at λem = 385 nm excitation at λexc = 471 nm
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HP
LC
pro
file
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HR
MS
(-)
ES
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1H
NM
R (
400 M
Hz,
D2O
, 25°C
)
m7GpCH2ppS_piren_D2O_1H_25deg.esp
8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5
Chemical Shift (ppm)
0
0.001
0.002
0.003
0.004
0.005
0.006
0.007
Norm
alized Inte
nsity
1.863.011.001.081.031.081.110.470.970.910.970.950.900.960.970.910.920.880.88
8.3
48.2
98.1
08.0
88.0
3
7.9
27.8
07.7
5 7.7
07.6
8
4.9
14.7
54.7
44.7
34.7
34.7
34.7
34.7
24.7
14.7
04.7
04.7
04.6
94.6
7 4.3
24.1
5 4.1
34.0
83.9
73.9
63.8
33.7
63.7
5
3.1
8
2.4
62.4
42.4
12.3
7
H1’
H5’’
H5’
H4’
H3’
H2’
-CH2-
-CH3 Water
aromatic (H(N1), H2-10)
-CH2 (phosph. bridge)
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31P
NM
R (
162 M
Hz,
D2O
, 25°C
)
m7GpCH2ppS_piren_31P_D2O_25deg
19 18 17 16 15 14 13 12 11 10 9 8 7 6 5
Chemical Shift (ppm)
-0.1
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
Norm
alized Inte
nsity
1.851.00
16.9
6
7.5
47.3
3
P α
Pβ, γ
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GDPβS-pyrene/NH4+ (2a)
F
orm
ula
Sp
ectr
osc
op
ic p
rop
erti
es
absorbance emission λexc = 282 nm emission λexc = 369 nm excitation at λem = 385 nm excitation at λexc = 471 nm
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HP
LC
rea
ctio
n p
rofi
le
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HP
LC
pro
file
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HR
MS
(-)
ES
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1H
NM
R (
400 M
Hz,
D2O
, 25°C
)
GDPbetaSPyrene_D2O_1H_25deg_RK.esp
8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0
Chemical Shift (ppm)
0
0.005
0.010
0.015
0.020
0.025
Norm
alized Inte
nsity
1.131.172.311.152.271.301.082.101.022.080.190.891.071.000.99
8.1
4 8.1
18.0
07.9
97.9
27.8
77.7
8
7.6
6 7.5
37.5
17.4
77.4
5
4.7
74.7
14.7
14.5
44.5
24.5
04.4
84.4
64.4
4
4.3
4
4.1
14.0
24.0
0
3.7
1
H1’
H5’, H5”
H4’
H3’
H2’
Water
aromatic (H(N1), H2-10)
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31P
NM
R (
162 M
Hz,
D2O
, 25°C
)
GDPbetaSPyrene_31P_D2O_25deg.esp
12 10 8 6 4 2 0 -2 -4 -6 -8 -10 -12 -14 -16
Chemical Shift (ppm)
-0.1
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
Norm
alized Inte
nsity
1.000.96
7.2
57.0
9
-11.7
5-1
1.9
5
Pα
Pβ
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GTPγS-pyrene/NH4+ (2b)
Form
ula
Sp
ectr
osc
op
ic p
rop
erti
es
absorbance emission λexc = 282 nm emission λexc = 369 nm excitation at λem = 385 nm excitation at λexc = 471 nm
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HP
LC
pro
file
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HR
MS
(-)
ES
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1H
NM
R (
400 M
Hz,
D2O
, 25°C
)
GTPgammaS_piren_D2O_1H_25deg.esp
8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0
Chemical Shift (ppm)
0
0.005
0.010
0.015
Norm
alized Inte
nsity
1.161.152.411.122.331.091.092.221.111.090.211.921.121.051.00
8.3
38.3
1
8.0
8
8.0
6 8.0
07.9
3 7.9
37.8
47.7
87.7
5 7.5
97.5
77.4
3
4.8
34.8
2
4.7
24.6
64.6
4
4.3
5 4.3
2
4.1
24.0
44.0
3
3.7
43.7
33.7
2
H1’
H5”
H4’, H5’
H3’
H2’
Water
aromatic (H(N1), H2-10)
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31P
NM
R (
16
2 M
Hz,
D2O
, 25°C
)
GTPgammaS_piren_31P_D2O_25deg.esp
5 0 -5 -10 -15 -20 -25
Chemical Shift (ppm)
-0.1
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
Norm
alized Inte
nsity
1.001.181.04
7.9
87.8
2
-11.1
4-1
1.2
5
-23.2
0 -23.3
2-2
3.3
7-2
3.4
8
Pγ
Pα
Pβ
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PS-pyrene/NH4+ (3)
Form
ula
Sp
ectr
osc
op
ic p
rop
erti
es
absorbance emission λexc = 282 nm emission λexc = 369 nm excitation at λem = 385 nm excitation at λexc = 471 nm
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HR
MS
(-)
ES
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1H
NM
R (
40
0 M
Hz,
DM
SO
-d6, 25°C
) PROTON_01_2.esp
8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0
Chemical Shift (ppm)
0
0.05
0.10
0.15
0.20
Norm
alized Inte
nsity
2.781.181.092.632.771.061.00
DMSO
8.8
08.7
78.6
88.6
68.3
98.3
7
8.3
18.3
08.2
88.2
38.1
38.1
1
4.3
04.2
7
2.5
12.5
02.5
02.5
02.4
9
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31P
NM
R (
162 M
Hz,
D2O
, 25°C
)
PHOSPHORUS_01
11.0 10.5 10.0 9.5
Chemical Shift (ppm)
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0N
orm
alized Inte
nsity
1.00
10.4
5
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gD
QC
OS
Y (
400 M
Hz,
D2O
, 25°C
)
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gH
SQ
CA
D (
400 M
Hz,
D2O
, 25°C
)
