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Kinetics of charge transfer in DNA containing a mismatch.

Osakada Y, Kawai K, Fujitsuka M, Majima T - Nucleic Acids Res. (2008)

Bottom Line: While the single-base mismatch would significantly affect the CT in DNA, the kinetic basis for the drastic decrease in the CT efficiency through DNA containing mismatches still remains unclear.We assumed that further elucidating of the kinetics in mismatched sequences can lead to the discrimination of the DNA single-base mismatch based on the kinetics.In this study, we investigated the detailed kinetics of the CT through DNA containing mismatches and tried to discriminate a mismatch sequence based on the kinetics of the CT in DNA containing a mismatch.

View Article: PubMed Central - PubMed

Affiliation: The Institute of Scientific and Industrial Research, Osaka University, Mihogaoka 8-1, Ibaraki Osaka 567-0047, Japan.

ABSTRACT
Charge transfer (CT) in DNA offers a unique approach for the detection of a single-base mismatch in a DNA molecule. While the single-base mismatch would significantly affect the CT in DNA, the kinetic basis for the drastic decrease in the CT efficiency through DNA containing mismatches still remains unclear. Recently, we determined the rate constants of the CT through the fully matched DNA, and we can now estimate the CT rate constant for a certain fully matched sequence. We assumed that further elucidating of the kinetics in mismatched sequences can lead to the discrimination of the DNA single-base mismatch based on the kinetics. In this study, we investigated the detailed kinetics of the CT through DNA containing mismatches and tried to discriminate a mismatch sequence based on the kinetics of the CT in DNA containing a mismatch.

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Bar graphs showing rate constants for CT in DNA containing a mismatch (kht). Rate constants of assembly-1 (a), assembly-2 (b) and assembly-3 (c), respectively. kht for fully matched sequences, G-containing mismatch and A/A, T/T or A/C are shown in gray, orange and green, respectively.
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Figure 4: Bar graphs showing rate constants for CT in DNA containing a mismatch (kht). Rate constants of assembly-1 (a), assembly-2 (b) and assembly-3 (c), respectively. kht for fully matched sequences, G-containing mismatch and A/A, T/T or A/C are shown in gray, orange and green, respectively.

Mentions: We first examined the effects of the G-containing mismatches on the CT. Excitation of the NI site with a 355-nm laser results in the formation of the NI radical anion (NI•−) with a peak at 400 nm immediately after the laser flash which remained almost constant on the time scale of the present experiment (≈100 μs) due to the slow charge recombination rate between NI•− and G•+ across six A–T base pairs (Supplementary Figures S2, S3 and S4) (22). Absorptions at 520 nm assigned to PTZ•+ then emerged on the time scale of several microseconds for the fully matched sequences, GC-1 and GC-2, and on the time scale of several dozen microseconds for GC-3 (Figure 2), which was in good agreement with the results of the kinetic modeling using the CT values for the CT between the fully matched Gs reported in our previous studies (Supplementary Table S1) (22,23). It is seen that the kinetics of the CT between Gs, that is, the CT from the G to G-containing mismatch, is slowed by the introduction of a G/T or G/A mismatch (Figure 2). The detailed kinetics of the CT between Gs was obtained using the Matlab software as previously described (23). The represented profiles were obtained from the accumulation of 32 laser shots (Figure 2). The rate constants were obtained on the basis of three independent experiments and we determined the errors from these experiments. The kinetic modeling as shown in Figures 1 and 3 was used to determine the rate constant for the CT. Employing the kinetic modeling based on the assumption that the back CT from the mismatched G to G through the intervening A/T base pairs is slow compared to the rapid CT between the GC repeats, we determined the rate constants of the CT from G to the G-containing mismatch. As for the rate constants for the GC repeats, we utilized the previously reported value (k = 2.2 × 108/s) (23). The rate constants of the CT from G to the G-containing mismatch are listed in Table 1 and represented in Figure 4 as bar graphs.Figure 2.


Kinetics of charge transfer in DNA containing a mismatch.

Osakada Y, Kawai K, Fujitsuka M, Majima T - Nucleic Acids Res. (2008)

Bar graphs showing rate constants for CT in DNA containing a mismatch (kht). Rate constants of assembly-1 (a), assembly-2 (b) and assembly-3 (c), respectively. kht for fully matched sequences, G-containing mismatch and A/A, T/T or A/C are shown in gray, orange and green, respectively.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
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Figure 4: Bar graphs showing rate constants for CT in DNA containing a mismatch (kht). Rate constants of assembly-1 (a), assembly-2 (b) and assembly-3 (c), respectively. kht for fully matched sequences, G-containing mismatch and A/A, T/T or A/C are shown in gray, orange and green, respectively.
Mentions: We first examined the effects of the G-containing mismatches on the CT. Excitation of the NI site with a 355-nm laser results in the formation of the NI radical anion (NI•−) with a peak at 400 nm immediately after the laser flash which remained almost constant on the time scale of the present experiment (≈100 μs) due to the slow charge recombination rate between NI•− and G•+ across six A–T base pairs (Supplementary Figures S2, S3 and S4) (22). Absorptions at 520 nm assigned to PTZ•+ then emerged on the time scale of several microseconds for the fully matched sequences, GC-1 and GC-2, and on the time scale of several dozen microseconds for GC-3 (Figure 2), which was in good agreement with the results of the kinetic modeling using the CT values for the CT between the fully matched Gs reported in our previous studies (Supplementary Table S1) (22,23). It is seen that the kinetics of the CT between Gs, that is, the CT from the G to G-containing mismatch, is slowed by the introduction of a G/T or G/A mismatch (Figure 2). The detailed kinetics of the CT between Gs was obtained using the Matlab software as previously described (23). The represented profiles were obtained from the accumulation of 32 laser shots (Figure 2). The rate constants were obtained on the basis of three independent experiments and we determined the errors from these experiments. The kinetic modeling as shown in Figures 1 and 3 was used to determine the rate constant for the CT. Employing the kinetic modeling based on the assumption that the back CT from the mismatched G to G through the intervening A/T base pairs is slow compared to the rapid CT between the GC repeats, we determined the rate constants of the CT from G to the G-containing mismatch. As for the rate constants for the GC repeats, we utilized the previously reported value (k = 2.2 × 108/s) (23). The rate constants of the CT from G to the G-containing mismatch are listed in Table 1 and represented in Figure 4 as bar graphs.Figure 2.

Bottom Line: While the single-base mismatch would significantly affect the CT in DNA, the kinetic basis for the drastic decrease in the CT efficiency through DNA containing mismatches still remains unclear.We assumed that further elucidating of the kinetics in mismatched sequences can lead to the discrimination of the DNA single-base mismatch based on the kinetics.In this study, we investigated the detailed kinetics of the CT through DNA containing mismatches and tried to discriminate a mismatch sequence based on the kinetics of the CT in DNA containing a mismatch.

View Article: PubMed Central - PubMed

Affiliation: The Institute of Scientific and Industrial Research, Osaka University, Mihogaoka 8-1, Ibaraki Osaka 567-0047, Japan.

ABSTRACT
Charge transfer (CT) in DNA offers a unique approach for the detection of a single-base mismatch in a DNA molecule. While the single-base mismatch would significantly affect the CT in DNA, the kinetic basis for the drastic decrease in the CT efficiency through DNA containing mismatches still remains unclear. Recently, we determined the rate constants of the CT through the fully matched DNA, and we can now estimate the CT rate constant for a certain fully matched sequence. We assumed that further elucidating of the kinetics in mismatched sequences can lead to the discrimination of the DNA single-base mismatch based on the kinetics. In this study, we investigated the detailed kinetics of the CT through DNA containing mismatches and tried to discriminate a mismatch sequence based on the kinetics of the CT in DNA containing a mismatch.

Show MeSH