Supporting Information
A Ru
thenium
(II) Complex as a
Luminescent
Probe for
DNA Mismatches and Abasic Sites
Adam N. Boynton, Lionel Marcélis, Anna J. McConnell, and Jacqueline K. Barton*
Division of Chemistry and Chemical Engineering
California
Institute of Technology, Pasadena, California, 91125
Figure S1:
UV
-
Visible spectrum of [Ru(bpy)
2
(
BNIQ
)
]
2+
(
6
μ
M
in H
2
O).
Figure S2:
Steady
-
state emission spectra of
[Ru(bpy)
2
(
BNIQ
)
]
2+
in aerated (solid line) and
deoxygenated
(
via
argon bubbling,
dotted line) solutions
(6
μ
M in H
2
O
,
l
ex
= 440 nm
).
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
0.5
215
265
315
365
415
465
515
565
Absorbance2
Wavelength2(nm)
0
5
10
15
20
25
30
600
650
700
750
800
850
Emission/Intensity/(
a.u
.)
Wavelength/(nm)
Figure S
3
:
Steady
-
state luminescence titrations of [Ru(bpy)
2
(
BNIQ
)
]
2+
with
well
-
matched
(blue), mismatched (red), and abasic (pink)
DNA
.
Samples
were
prepared in 5 mM
T
ris,
200
mM NaCl, pH 7.5.
[Ru] = 4
μ
M,
l
ex
= 440 nm.
[DNA] reflects the concentration of full
sequence.
Emission
spectra were integrated from 590
-
850
nm.
Binding affinities
of the complex for well
-
matched
,
mismatched
, and abasic
sites
were
evaluated by a global f
itting process using
a
modified
McGhee
-
Von Hippel method
.
1
Briefly, we
consider the
equilibrium binding
(K
ass
)
of the complex (
C
)
to well
-
matched base pair sites (BP)
and the equilibrium binding (K
MM
) to a mismatch
(or abasic)
site (MM):
퐶
+
퐵푃
⇌
[
퐶
_
퐵푃
]
퐾
+,,
=
[
.
_
/0
]
.
/0
퐶
+
푀푀
⇌
[
퐶
_
푀푀
]
퐾
22
=
[
.
_
22
]
.
22
As described previously,
2
we can express the luminescence intensity
(
I
) as a function of these
equilibria along with other parameters, defined below.
1
1.1
1.2
1.3
1.4
1.5
1.6
1.7
1.8
0
0.5
1
1.5
2
2.5
3
I/I
0
[DNA]/[Ru]
WM
MM
AB
퐼
=
훼
퐾
+,,
퐶
.
푛
1
−
푥
푅
푓
1
+
퐾
+,,
퐶
.
푝
푓
+
훽
퐾
22
퐶
.
푛
푥
푅
푓
1
+
퐾
+,,
퐶
.
푓
In this equation for
I,
C
c
is the total concentration of complex;
R
is the ratio of total DNA duplex
concentration to
C
c
;
f
is the molar fraction of free complex
;
n
is the number of
base pairs per
DNA duplex;
x
is the ratio of mismatched
to well
-
matched
sites in the duplex;
and
p
is the
occupational factor
.
Important to note are
훼
and
훽
, which
represent the
the
emissivities of the
complex when bound to a well
-
matched or mismatched site, respectively, relative to free
complex.
The
global fitting on the
three
data
set
s
is performed (occupational factor set to 2) and
yield
s
the values of K
a
=
7.3
10
3
M
-
1
per
well
-
m
atched
base pair
,
K
a
=
3
.
5
10
6
M
-
1
per
CC
mismatch site,
and
K
a
=
3
.
8
10
6
M
-
1
per
abasic site. The
emissivities
for
the
complex associated
with these site
s
, relative to the lumi
nescence of the free complex
,
are estimated to
be
1.36, 1.42
and 1.46 for
well
-
matched
,
mismatched
,
and abasic site
s,
respectively
. The errors are evaluated
to be equal to 10%.
Figure S4:
Ferricyanide quenching of
free
[Ru(bpy)
2
(BNIQ)]
2+
(black) and in the presence of
well
-
matched DNA (blue) and mismatched DNA (red).
Ferricyanide was added (dotted lines) to
concentrations of 1.2, 2.3, 3.5, and 5.6 mM.
[Ru] = 2
μ
M
, [DNA] = 4
μ
M
,
λ
ex
= 440 nm.
Samples
were
prepared in 5 mM Tris, 200 mM NaCl, pH 7.5 at 25
°C.
Supporting
References
1.
McGhee, J.D.; von Hippel, P.H.
Theoretical Aspects of DNA
-
Protein Interactions: Co
-
operative and Non
-
co
-
operative Binding of Large Ligands to a One
-
dimensional
Homogeneous Lattice.
J. Mol. Biol
.
1974
,
86
, 469
-
489.
2.
Boynton, A.N.; Marcelis, L.; Barton, J.K.
[Ru(Me
4
phen)
2
(dppz)]
2+
, A Light Switch for
DNA Mismatches.
J. Am. Chem. Soc.
2016
,
138
, 5020
-
5023.
0
2
4
6
8
10
12
600
650
700
750
800
850
Emission/Intensity/(a.u.)
Wavelength/(nm)
0
1
2
3
4
5
6
7
8
600
650
700
750
800
850
Emission/Intensity/(a.u.)
Wavelength/(nm)
0
1
2
3
4
5
6
7
8
600
650
700
750
800
850
Emission/Intensity/(a.u.)
Wavelength/(nm)
Inc.%
[Fe]
Inc.%
[Fe]
Inc.%
[Fe]
Ru
WM
MM