OTHER INSTRUMENTAL METHODS 141
oxide - sharpened nanoprobes), and (4) compatible substrates (such as salinized
mica and carbon coated mica). The ability to generate nanometer - resolved
images of unmodifi ed nucleic acids has broad biological applications. Chromo-
some mapping, transcription, translation, and small molecule – DNA interac-
tions such as intercalating mutagens provide exciting topics for high - resolution
studies. For example, one study looked at the effect of Ni(II) ions on double -
stranded, ds, GC - DNA (DNA strands having only guanine and cytosine base
pairs, dG – dC · dG – dC) versus the effect on ds AT - DNA (DNA strands having
only adenine and thymine base pairs, dA – dT · dA – dT).^45 It is known that many
metal ions affect DNA structure, especially a B - to Z - DNA structural trans-
formation in GC - DNA. Group I and II metal cations — Na + , K + , Ca 2+ , Mg 2+ —
and, at much lower concentrations, transition metal divalent cations — Ni(II),
Co(II), Zn(II) — are known to cause the B - to Z - DNA transformation. The
reference 45 researchers used AFM to detect condensed forms of the two
DNA types in the presence of NiCl 2 solutions of differing molarities. They
detected condensed forms of GC - DNA — toroids, rods, and jumbles — in solu-
tions with 0.5 M Ni(II) concentrations, whereas AT - DNA only began to form
condensed structures at 6 M concentrations of Ni(II). The researchers con-
cluded that the better ability of GC - DNA to form condensed structures was
due to the formation of Ni(II) – N7 coordination with major - groove available
guanine base N7 positions. In contrast, Ni(II) interaction with AT - DNA
appeared to occur only electrostatically in the DNA minor groove and not
with coordination positions on the adenine or thymine bases. It is known that
the guanine base N7 position is the one most favored by transition metals in
DNA binding as is well known for the anticancer drug, cis - diamminedichloro -
platinum(II), cisDDP (see Figure 1.6 ). The reference 45 researchers also used
the “ electrostatic zipper model ” for DNA aggregation to show that GC - DNA
was more likely to “ zip ” itself into agglomerated structures than was the AT -
DNA molecule. They concluded that AFM was an excellent technique for
detecting condensed structures in dsDNA.
Researchers have also succeeded in imaging individual proteins and other
small molecules with the AFM. Smaller molecules that do not have a high
affi nity for common AFM substrates have been successfully imaged by employ-
ing selective affi nity binding procedures. Thiol incorporation at both the 5 ′ and
3 ′ ends of short PCR products (PCR, polymerase chain reaction, described in
Section 2.3.5) has been shown to confer a high affi nity for ultrafl at gold sub-
strates and therefore improved AFM imaging. Small proteins, like cytochrome
c (see Section 7.7) , have been imaged interacting with their redox partners.
For example, Choi and Dimitriadis have used AFM to detect how cytochrome
c binds to the lipids in the mitochondrial inner membrane.^46 As discussed in
much more detail in Chapter 7 , cytochrome c plays an essential role in the
respiratory electron transport chain by shuttling electrons between cytochrome
c reductase and cytochrome c oxidase. Also, release of cytochrome c from lipid
interactions in the mitochondrial inner membrane is believed to be important
fi rst step in cell death (apoptosis). Experimentally, the researchers used AFM