Selective inhibition of DNA
replication by ph–PNA conjugates. (A)
An in vitro replication run-off assay from a mtDNA
template containing the MERRF mutation was performed. In the absence
of ph–PNA this gave a product of 341 nt. In the presence
of 50 or 100-fold molar excess of ph–PNA a truncated product of
245 nt was formed. (B) An in vitro replication
run-off assay from a wild-type mtDNA template was performed. This
template differed from the MERRF template in (A) at a single nucleotide
position in the PNA binding region. In the absence of ph–PNA
this gave a full-length product of 350 nt (the MERRF DNA template is
shorter due to a 9 bp deletion at the 3′ end,
downstream of the replication inhibition site). Even in the presence
of a 500-fold molar excess of ph–PNA none of the truncated
product of 245 nt was found, in contrast to the positive control
using the MERRF template. The binding affinities of PNA and ph–PNA
to the MERRF PNA templates were 11.3 ± 2.2
and 37 ± 6 nM, respectively (Paul Smith,
University of Newcastle upon Tyne, personal communication).