Biomedical Engineering Reference
In-Depth Information
a demonstration that two proteins, azurin and mdm2, do not compete for the same
binding site on p53.
6.3.4 DFS S
TUDIES OF
C
OMPLEXES
I
NVOLVING
DNA
OR
A
PTAMERS
Deoxyribonucleic acid and ribonucleic acid (RNA) molecules drive and regulate
the storage, transport, processing, and expression of genetic information in living
organisms. DNA consists of two long polymers of nucleotides organized in com-
plementary strands kept together by noncovalent bonds. RNA, which is derived from
DNA through transcription process, differs from DNA by the substitution of the sugar
deoxyribose with ribose, and it is usually single stranded. Since gene expression in
eukaryotes is controlled by the specific binding of transcription factors to defined
DNA sequences, the possibility to influence and control cell functions through
modified synthetic transcriptional factors at single-molecule level offers fascinating
prospects for molecular biology. Recently, a new class of molecules derived from
DNA, called
aptamers
, have received a large attention, especially for applicative
purposes (Cho et al., 2009). Aptamers are synthetic oligonucleotides derived from
in vitro
experiments in which single-stranded DNA or RNA molecules are isolated
from a random sequence library according to their ability to selectively bind target
molecules, which may be organic dyes, drugs, metal ions, amino acids, peptides, and
proteins (Ellington and Szostak, 1990). With respect to antibodies, which, indeed, are
widely used, aptamers present many advantages, including simpler synthesis, faster
tissue penetration, easier modification and storage, and wider applicability; therefore,
they may strongly rival antibodies in both diagnostic and therapeutic applications.
Table 6.4 reports the results from some DFS studies on (1) DNA molecules;
(2) complexes involving DNA in interaction with peptides, proteins, drugs, and so
on; and (3) aptamer-base complexes. It is interesting to remark that the unbind-
ing force values for these systems are similar to those measured for protein pairs
(see Tables 6.1 through 6.3). This suggests that the noncovalent interactions regu-
lating biomolecular association possess general features, irrespective of the specific
molecules involved.
The first DFS studies on DNA have been fully devoted to investigate the inter-
action between complementary DNA strands. With such an aim, complementary
single strands of a DNA have been immobilized on an AFM tip and a surface via
linkers (see Figure 6.9). Starting from the preliminary results of a previous work
(Lee et al., 1994a), Strunz et al. have compared the unbinding force between DNA
strands formed by 10, 20, and 30 base pairs (bp) (Strunz et al., 1999). For all these
systems, the unbinding force plotted as a function of the logarithm of the loading
rate has revealed a linear regime indicative of the existence of a single energy barrier
to be overcome to break the double DNA strand. Accordingly, the kinetic mecha-
nism regulating such a process has been described as a thermally driven dissoci-
ation, similarly to what happens for protein-protein interactions. In addition, they
found that the width of the energy barrier linearly increases with the number of bp
while the dissociation rate exhibits an exponential decrease. These effects have been
interpreted in terms of a cooperative unbinding effect of the bp responsible for the
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