Chemistry Reference
In-Depth Information
chromophore's excited state dynamics. Characteristic changes to DNA are observed
for photobinding of metal complexes, photooxidation of nucleobases and photo-
cleavage of the sugar phosphate backbone. Knowledge of the decomposition path-
ways of DNA photolyzed with a metal complex helps to understand a possible
mechanism of PDT action.
8.3.1 DNA Targets and Analysis
Study of the photochemical reactions of metal complexes with DNA requires careful
choice of the type of DNA used and method of analysis. Helpful descriptions of
DNA structure and analysis can be found in the topic by Calladine.
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Mono - and
oligonucleotides are used as targets when chemical analysis techniques such as mass
spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy are used
to study the photoproducts. Mononucleotides are available commercially and have
found utility in experiments to understand binding of metal complexes to DNA and
ES quenching studies. Oligonucleotides (oligos) are short strands of DNA (15- 100
base pairs) that have known sequence and structural properties. Oligos are better
models for nuclear DNA because they can be obtained as double stranded DNA.
High-resolution poly(acrylamide) gel electrophoresis (HR-PAGE) is instrumental
in understanding sequence specifi city of photochemical reactions with oligonuclear
DNA leading to irreversible changes. Circular plasmid DNA extracted from bacte-
ria is obtained with a supercoiled tertiary structure that makes this type of DNA an
excellent target for photomodifi cation schemes.
54,55
Changes to the topographical
nature of supercoiled circular plasmid DNA are indicative of several decomposition
pathways that can be analysed using agarose gel electrophoresis (Figure 8.17).
Finally, genomic DNA is commercially available as a freeze dried extract from
various tissue sources and is high molecular weight, with
10
6
base pairs per mole-
cule. Genomic DNA is used to understand DNA-metal interactions that require
large volumes and/or high DNA concentrations for analysis using selective precipi-
>
λ
Cir Lin
23.1 kbp-
9.4 kbp-
6.6 kbp-
4.4 kbp-
Form II
Form III
2.3 kbp-
2.0 kbp-
Form I
Figure 8.17
Photograph of a typical 0.8% agarose gel illustrating the electrophoretic migra-
tion (moving from top to bottom) of three different forms of plasmid DNA. The lanes from
left to right are as follows:
l
is a lambda DNA/HindIII digest molecular weight marker
(kbp = 1,000 base-pairs), Cir is circular plasmid DNA containing native supercoiled (Form I)
and open-circular (Form II) plasmid DNA, and Lin is linear plasmid DNA (Form III). Band
assignments are based on Vinograd and Lebowitz
55
