Supplementary Material to:
More pronounced salt dependence and higher reactivity for
platination of the hairpin r(CGCGUUGUUCGCG) compared with
d(CGCGTTGTTCGCG)
Margareta Hägerlöf1, Pal Papsai1, Christine S. Chow2, Sofi K. C. Elmroth1,*
10
1
Biochemistry, Chemical Center, Lund University, P.O. Box 124, SE-221 00 LUND,
Sweden, and 2Wayne State University, Department of Chemistry, Detroit, MI 48202,
USA
Table S1 Observed first-order- and apparent second-order rate constants determined
by use of HPLC technique for platination of RNAI and DNAI by complexes 1, 2, and
3 in buffered solution at pH 6.0; 5.0 mM phosphate buffer for [Na+] in the interval 19
mM ≤ I ≤ 500 mM, and 0.50 mM phosphate buffer for[Na+] = 1.0 mM.
Fig. S1 A HPLC traces at different reaction times for the reaction of RNAI and 2 in
20
5 mM Na-phosphate buffer pH 6.0, [NaClO4]= 95 mM, T = 25 °C, [RNAI] = 1.25
M, CPt = 74 M. The insert shows a plot of normalized HPLC peak areas
corresponding to the peak labeled as () as a function of reaction time.
B HPLC traces at different reaction times for the reaction of DNAI and 3 in 5 mM
Na-phosphate buffer pH 6.0, 295 mM NaClO4, T = 25 °C, 1.25 M DNAI, CPt = 74.2
M. The insert shows a plot of normalized HPLC peak areas corresponding to the
peak labeled as () as a function of reaction time.
Fig. S2 Observed pseudo-first-order rate constants as a function of increasing CPt for
the reaction of RNAI and 2. Reaction conditions: [RNAI] = 1.25 M, 5.0 mM Na-
30
phosphate buffer, pH 6.0, 30 mM NaClO4, 50 M < CPt < 150  M (40- to 120-fold
excess).
Fig. S3 log(k2app) versus √I/(√I+1) as monitored for the reaction between platinum
complexes 2 and 3 with RNAI and DNAI at 25 °C, pH 6.0, 19 mM ≤ [Na+] ≤ 500
mM, [Pt(II)] = 7.5  10-5 M, [oligonucleotide] = 1.25  10-6 M. Reaction of RNAI and
2 (), reaction of RNAI and 3 (), reaction of DNAI and 2 (), reaction of DNAI
and 3 ().
Fig. S4 A Autoradiogram illustrating rough reaction kinetics resulting from
incubation of complex 3 with 3´-end labeled RNAI; CRNA = 2.2 M CPt = 120  M.
The reactions were studied in Buffer A at 25 °C and were analyzed at time intervals; 0
10
min (lane 1), 1 min (lane 2), 6 min (lane 3), 20 min (lane 4), 40 min (lane 5), 1 h (lane
6), 1.5 h (lane 7), 2.5 h (lane 8), 4 h (lane 9), 6.75 h (lane 10) (20% denaturing
PAGE) B Autoradiogram illustrating rough reaction kinetics resulting from
incubation of complex 3 with 3´-end labeled RNAI; CRNA = 2.2 M, CPt = 220 M.
The reactions were studied in Buffer A at 25 °C and were analyzed at time intervals; 0
min (lane 1), 1 min (lane 2), 6 min (lane 3), 15 min (lane 4), 36 min (lane 5), 1.1 h
(lane 6), 1.5 h (lane 7), 2 h (lane 8), 3 h (lane 9), 5.5 h (lane 10) (20% denaturing
PAGE) C Autoradiogram illustrating rough reaction kinetics resulting from
incubation of complex 3 with 5´-end labeled DNAI; CDNA = 2.07 M CPt = 87 M.
The reactions were studied in Buffer A at 37 °C and were analyzed at time intervals; 0
20
min (lane 1), 1 min (lane 2), 6 min (lane 3), 15 min (lane 4), 30 min (lane 5), 50 h
(lane 6), 1h 6 min (lane 7), 1.5 h (lane 8), 2 h (lane 9), 3 h (lane 10), 4 h (lane 11)
(20% denaturing PAGE).
Fig. S5 A Thermodynamic data for the melt of RNAI in Buffer B (195 mM NaCl, 5
mM phosphate buffer pH 6.0, [Na+] = 200 mM), absorbance versus temperature
profiles for RNA concentrations1028
M, 359
M, and 136
M B
Thermodynamic data for the melt of RNAI-1 in Buffer B (195 mM NaCl, 5 mM
phosphate buffer pH 6.0, [Na+] = 200 mM), absorbance versus temperature profiles
for RNAI-1 concentrations 75
30
M and 9.9
M.
Table S1 Observed first-order- and apparent second-order rate constants determined
by use of HPLC technique for platination of RNAI and DNAI by complexes 1, 2, and
3 in buffered solution at pH 6.0; 5.0 mM phosphate buffer for [Na+] in the interval 19
mM ≤ I ≤ 500 mM, and 0.50 mM phosphate buffer for[Na+] = 1.0 mM.
[Na+] / 10-3M
kobs / 10-4 s-1
k2,app / M-1s-1
1
35
4.4 ± 0.4
5.9 ± 0.5
RNAI
2
1
21.8 ± 1.1
29.0 ± 1.5
RNAI
2
35
4.0 ± 0.4
5.3 ± 0.6
RNAI
2
75
2.29 ± 0.19
3.05 ± 0.25
RNAI
2
100
2.02 ± 0.21
2.69 ± 0.28
RNAI
2
300
0.96 ± 0.04
1.28 ± 0.05
RNAI
2
500
0.717 ± 0.014
0.956 ± 0.018
RNAI
3
35
59.2 ± 1.3
79.0 ± 1.7
RNAI
3
75
43.275 ± 0.5
57.7 ± 0.7
RNAI
3
100
37.8 ± 3
50.4 ± 4
RNAI
3
500
20.3 ± 2.4
27 ± 3.2
DNAI
2
1
12.1 ± 1.1
16.1 ± 1.5
DNAI
2
19
4.6 ± 0.5
6.1 ± 0.7
DNAI
2
35
3.5 ± 0.3
4.7 ± 0.4
DNAI
2
100
1.77 ± 0.13
2.36 ± 0.17
DNAI
2
300
1.6 ± 0.3
2.1 ± 0.4
DNAI
3
1
74 ± 6
99 ± 9
DNAI
3
19
30 ± 2.1
40.3 ± 2.9
DNAI
3
35
21.9 ± 0.8
29.2 ± 1.0
DNAI
3
75
10.4 ± 0.3
13.9 ± 0.4
DNAI
3
100
9.0 ± 0.6
12.0 ± 0.8
DNAI
3
300
6.9 ± 0.3
9.2 ± 0.4
DNAI
3
500
6.4 ± 0.6
8.6 ± 0.8
Target
Platination
oligo
reagent
RNAI