Chemistry Reference
In-Depth Information
7.4.3.2
Formation of “Hot Spots“ for Hydrolytic Scission at a Predetermined Position
These findings can be extended to “non-covalent“ site-selective DNA hydrolysis [13].
With single-stranded DNA as substrate, the additives used are two oligonucleotides
that are complementary with part of the substrate DNA. Upon mixing both these ad-
ditives with substrate DNA, most of the DNA forms duplexes with the oligonucleotide
additives and only the target site is kept single-stranded (this kind of structure is called
as a gap). The position of the gap and its size (number of unpaired nucleotides) depend
of course on the oligonucleotide additives and, thus, are easily controllable. These gap-
sites are “hot spots“ for catalysis by Ce(
IV
)/EDTA and are selectively hydrolyzed [Figure
7.1(b)]. Conversely, double-stranded portions in the substrate DNA are kept intact be-
cause of the poor reactivity. Covalent immobilization of the Ce(
IV
) complex to the target
site is unnecessary.
Alternative “hot spots“ are bulge-sites in substrate DNA, formed by using an oligo-
nucleotide additive that is several nucleotides shorter than the DNA substrate [13]. To
make “hot spots“ in double-stranded DNA substrates, invasion of peptide nucleic acid
(PNA) is quite useful, as detailed in Section 7.6.
7.5
Site-selective Scission of Single-stranded DNA
7.5.1
Promotion of Gap-selective DNA Hydrolysis by Introducing Monophosphate Groups to
the Gap-site
As described above [Figure 7.1(b)], gap-sites in single-stranded DNA substrate, formed
by “unmodified“ oligonucleotides, can be selectively hydrolyzed by Ce(
IV
)/EDTA com-
plex. However, both scission efficiency and site-selectivity are not satisfactorily high
for practical applications. To promote these two factors, monophosphate groups are
introduced to both edges (or either of them) of the gap-site [14]. Both scission effi-
ciency and site-selectivity are remarkably improved. The monophosphate groups
bind the Ce(
IV
) complex and recruit it to the target site (Figure 7.2).
Figure 7.2 Promotion of gap-selective DNA hydrolysis by introducing monophosphate
groups to the gap-site.
