S
1
Suppo
rting
Infor
mation
Insertion of a Bulky Rhodium Complex
into a DNA Cytosine
-
Cytosine Mismatch:
An NMR Solution Study
Christine Cordier
†
, Valérie C. Pierre
§
,
Jacque
line K. Barton
*
Division
of Chemistry and Chemical Engi
neering,
California Institute
of Technol
ogy,
Pasadena, California 91125,
USA
* to whom
corresponde
nce shoul
d be addr
essed at
jkba
rton@
caltech.edu
†
Current addr
ess:
ITODYS, UMR CNRS 7086,
Université Denis Diderot, Paris VII, 1 rue Guy
de
la
Brosse, 75005
Paris
, France.
§
Current address: Department of Chemistry, University of Minne
sota,
207
Pleasant St., SE
,
Minne
apol
is, MN 55455
, USA.
S
2
Tab
le S1.
NOE cont
acts of the free oligonuc
leotide d(CGGACTCCG). All chemical shifts are relative to
D
SS
-
d
6
(
δ
= 0.000
ppm
). The chemical shifts of the non
-
exchange
able and
exchange
able protons
were
measured at 10
°C
and
4 °C
respectively. Expe
rimental condi
tions
: [dsDNA] = 2.32
mM, 50
mM Pi, 20
mM NaCl, pH
= 6.10(
2).
Residue
H6/H8
H1’
H5/H2/Me
H2’
H2”
H3’
H4’
H
5’
H5”
NH / NH
2
C1
7.615
5.758
5.929
1.852
2.352
4.692
4.062
3.962
3.705
8.334
-
7.245
G2
7.921
5.477
-
2.676
2.698
4.707
4.304
4.072
3.962
13.
292
G3
7.799
5.648
-
2.647
2.755
5.037
4.414
4.194
4.133
12.
804
A4
8.182
6.259
7.957
2.749
2.829
4.954
4.215
4.
499
4.414
-
C5
7.261
5.721
5.404
2.431
1.772
4.695
4.102
4.365
4.047
-
T6
7.615
6.051
1.689
2.369
2.534
4.891
4.238
?
4.047
14.
406
C7
7.603
6.002
5.721
2.153
2.448
4.823
4.175
4.108
?
8.481
-
6.961
C8
7.542
5.672
5.734
2.034
2.369
4.854
4.141
4.108
4.072
8.823
-
7.210
G9
8.019
6.222
-
2.681
2.386
4.719
4.210
4.145
4.096
13.
230
S
3
Tab
le S2.
NOE conne
ctivities of the Rh
-
bound
oligonuc
leotide. All chemical shifts are relative to DSS
-
d
6
(
δ
= 0.000
ppm
).
The chemical shifts of the non
-
exchange
able and
exchange
a
ble protons
were measured
at 10
°C
and
4 °C
respectively. The two nO
e walks resulting
from
the loss of the C
2
sym
metry in the
central part of the oligonuc
leotide are indi
cated as a (blue) and
b (green). For A
4
, onl
y the amino
protons
of the a and
b strands
were distingui
shable. Expe
rimental condi
tions
: [dsDNA] = 1.62
mM, [
Δ
-
Rh(bpy
-
d
8
)
2
chrysi
3+
] = 1.62
mM (1 equi
valent per mismatch),
50
mM Pi, 20
mM NaCl, pH
= 6.10(
2).
§
A
mbiguous
assignm
ent.
Line broadening
and
ove
rlaps rende
r the distinction
between H5’ and
H5”
unc
ertain.
Residue
H6/H8
H1’
H5/H2/Me
H2’
H2”
H3’
H4’
H5’
/
H5”
NH / NH
2
C1
7.583
5.699
5.676
1.795
2.318
4.698
4.095
§
3.948
-
3.948
8.333
-
6.905
G2
7.908
5.441
-
2.619
2.619
5.022
4.192
4.192
-
4.026
13.
222
G3
7.791
5.555
-
2.622
2.642
5.051
4.351
4.130
-
4.026
13.
222
A4a
A4b
7.972
′′
6.017
′′
7.973
′′
2.210
′′
2.5
45
′′
4.857
′′
4.413
′′
4.114
-
3.974
′′
8.421
-
6.959
8.323
-
7.208
C5a
C5b
7.661
7.557
6.083
5.829
5.741
5.881
2.239
2.261
2.366
2.398
4.817
4.897
4.
4
13
4.660
4.049
-
3.952
4.017
-
3.978
-
-
T6a
T6b
7.615
7.511
5.744
5.679
1.946
1.940
2.199
2.053
2.369
2.364
4.
875
4.527
4.420
4.283
4.140
-
3.965
4.101
-
3.978
12.
793
12.
793
C7
7.557
5.715
5.786
2.097
2.443
4.840
4.156
4.101
-
?
8.743
-
7.227
C8
7.550
5.503
5.773
2.125
2.369
4.892
4.108
4.049
-
?
8.743
-
7.217
G9
7.973
6.147
-
2.704
2.368
4.694
4.192
4.121
-
4.065
12.
793
S
4
Tab
le S3.
Chemical shifts of the hydr
oge
ns of the chrysi ligand
in
the absence and
presence of
mismatched DNA.
All chemical shifts are relative to DSS
-
d
6
(
δ
= 0.000
ppm
). Expe
rimental condi
tions
:
[dsDNA] = 1.62 mM, [
Δ
-
Rh(bpy
-
d
8
)
2
chrysi
3+
] = 1.62
mM (1 equi
valent per mismatch),
50
mM Pi, 20
mM NaCl, pH
= 6.10(
2).
The chemical shifts of the non
-
exchange
able and
exchang
eable protons
were
measured at 10
°C
and
4
°C
respectively.
Proton
Unbound
Δ
-
Rh(bpy
-
d
8
)
2
chrysi
3+
DNA bound
by
Δ
-
Rh(bpy
-
d
8
)
2
chrysi
3+
χ
0
8.703
13.
032
χ
1
8.164
7.210
χ
2
7.516
7.018
χ
3
7.767
7.923
χ
4
8.242
6.188
χ
5
8.303
7.198
χ
6
8.303
7.934
χ
7
8.059
6.984
χ
8
7.515
7.241
χ
9
7.393
7.276
χ
10
7.889
7.364
χ
11
8.703
12.
900
S
5
Tab
le S4.
Intermolecular nO
e between
Δ
-
Rh(bpy
-
d
8
)
2
chrysi
3+
and
the oligonuc
leotide. The two strands
,
resulting
from
the loss of the C
2
sym
metry in the central part of the oligonuc
leotide, are marked a and
b.
Expe
rimental c
ondi
tions
: [dsDNA] = 1.62
mM, [
Δ
-
Rh(bpy
-
d
8
)
2
chrysi
3+
] = 1.62 mM (1 equi
valent per
mismatch),
50
mM Pi, 20
mM NaCl, pH
= 6.10(2), D
2
O, 10
°C
.
NOEs
from
a strand
NOEs
from
b strand
χ
1
–
T6aH2”
χ
7
–
C5bH
1’ w
χ
1
–
T6aH5’/H5”
χ
7
–
T6bM
e
χ
1
–
T6aMe
χ
8
–
C5bH
4
’
χ
2
–
C5aH1’
χ
8
–
C5bH
5
χ
2
–
C5aH4’
χ
8
–
T6bM
e
χ
2
–
T6aH2’
χ
9
–
C5bH
4’
χ
2
–
T6aMe
χ
9
–
T6bH
2”
χ
2
–
T6aH5’/H5”
χ
9
–
T6bM
e
χ
3
–
C5aH1’
χ
10
–
T6bi
χ
3
–
C5aH4’
χ
11
–
T6bi
χ
3
–
T6aMe
S
6
Figu
re S1
. Titration of
Δ
-
Rh(bpy
-
d
8
)
2
chrysi
3+
to (a) an oligon
ucleotide cont
aining
two CA mismatches
(d(CGATCGACCG), Tm = 13 ° C) and
(b) an oligonuc
leotide cont
aining
a singl
e CC mismatch
(d(CGGACTCCG), Tm = 17.8 ° C). For clarification,
onl
y the arom
atic region
is represented.
Expe
rimental condi
tions
: phos
pha
te buf
fer, I = 50 mM, T = 20
°C
, (a) pH
= 6.03, (b) pH
= 6.10.
S
7
Figu
re S2.
UV absorption at
λ
= 260
nm
of the free oligonuc
leotide (blue circle) and the metalloinsertor
-
bound
DNA (red squa
re).
The Tm values
represent the midpoi
nt of
the transition
as obt
ain
ed by fi
tting
the
melting
profi
les with a
sigm
oidal expr
ession.
S
8
Figu
re S3.
Phot
ocleavage induc
ed by the metalloinsertor. (a) MALDI
-
TOF mass spectrum
obt
ained after
1 hour
of irradiation
with a solar simulator. The produc
ts correspond
to cleavage at th
e T
6
neighbor
ing the
CC mismatch. (b) MALDI
-
TOF mass spectrum
obtained on
an identical sample without
phot
o
-
irradiation.
The M
1+
and M
2+
peaks correspond
to the unc
leaved 5’
-
CGGACTCCG
-
3’ oligonuc
leotide.
S
9
Figu
re S4.
2D
DQF
-
COSY sub
-
spectrum
of the free
oligonuc
leotide at
10
°C
(F2
×
F1 : H2’
-
H2”
×
H1’).
The crosspeak
patterns
indi
cate the conf
ormation
of the suga
rs. All suga
rs, includi
ng
that of the
mismatched cytosine, maintain the C
2
’
-
endo
puc
kering.
Inserts: t
he crosspeaks associated to the
A4 and
C5 s
uga
r
s are onl
y visible at a low
er signa
l
-
to
-
noi
se resolution.
S
10
Figu
re S5.
NOESY
sub
-
spectra and
assignm
ents of the nO
e cont
acts between the exchange
able protons
of the free oligonuc
leotide (uppe
r part:
F2
×
F1 : arom
atic + amino
×
arom
atic + amino;
low
er
part:
F2
×
F1 : arom
atic + amino
×
imino
)
.
The chemical shifts and
the crosspeaks of the imino
and
amino
protons
indi
cate that all bases are Watson
-
Crick
paired. No
correlation
is obs
erved
for the
C
-
C
mismatch,
sugge
sting
that the two cytosines are proba
b
ly paired by
a singl
e hyd
roge
n bond
in a Wobbl
e
-
type
conf
ormation.
NHb and
NHf correspond
to the bound
and
free amino
protons
respectively. Expe
rimental
condi
tions
: H
2
O, 4 °C
, 300
ms mixing
time.
S
11
Figu
re S6.
1D
1
H sub
-
spectra of the arom
atic, amino
and
imino protons
of (a) the free oligonuc
leotide
and
(b)
Δ
-
Rh(bpy
-
d
8
)
2
chrysi
3+
inserted in the DNA
-
Rh adduc
t. Expe
rimental condi
tions
, H
2
O, 4 °C
.
S
12
Figu
re S7.
NOESY
sub
-
spectra and
assignm
ents of the NOE cont
acts between the exchange
able protons
of the Rh
-
bo
und
DNA. (low
er part:
F2
×
F1 : aromatic + amino
×
imino;
medium
part:
F2
×
F1 : arom
atic
+ amino
×
arom
atic + amino;
uppe
r part: F2
×
F1 : imino
×
T6M
e groups
)
. NHb and
NHf correspond
to
the bound
and
free amino
protons
respectively. Loss of the
C
2
sym
met
ry in the central part of the
oligonuc
leotide results in two une
qui
valent strands
labeled a (blue) and
b (green). Intramolecular nO
e
correlations
between
chrysi protons
(labelled
χ
) are indi
cated
by
a pur
ple rectangl
e. Expe
rimental
condi
tions
: H
2
O, 4 °C
, 300
ms mixing
time.
S
13
Figu
re S8.
(a) COSY and
(b) NOESY sub
-
spectra and
assignm
ents of the arom
atic protons
of the chrysi
ligand
of unbound
Δ
-
Rh(bpy
-
d
8
)
2
chrysi
3+
(D
2
O, 20
°C
). In t
he COSY spectrum
two chains of scalar
correlations
match the two spin systems of the ligand.
* Weak correlations
obs
ervable onl
y at low
signa
l
-
to
-
noi
se resolution.