(1) the polar side chain must go from aqueous solvent through the hydrophobic region in between two adja-
cent base pairs, (2) the size and rigidity of the side chains may impede this passage, and (3) the rate of DNA
thermal motion will result in the formation of a bubble-like structure that can accommodate the side chain.
9.7 Interactions in the Minor Groove
9.7.1 General Characteristics of Groove Binding
A larger number of functional groups of the DNA bases are accessible in the wide major groove compared
to the narrow minor groove of the B-helix. Therefore, most proteins have evolved to make sequence-
specific interactions with DNA in the major groove. However, some proteins and many small molecules
interact with DNA in the minor groove. The major and minor grooves differ in electrostatic potential,
hydrogen-bonding characteristics, steric effects, and hydration. Furthermore, DNA structure and conform-
ation is highly dependent on sequence context. For example, runs of successive adenosine residues give
rise to a very narrow minor groove, where the base pairs have a higher propeller twist and the DNA
becomes slightly bent from the helical axis. In general, small groove-binding molecules exhibit a prefer-
ence for the minor groove, not least because this site of interaction provides better van der Waals contacts.
In addition, many minor groove-binding ligands prefer A T sites (compared to intercalators, which gener-
ally exhibit a G C preference).
Minor groove-binding ligands often contain several simple aromatic rings, such as pyrrole, furan, ben-
zeneor imidazole. These are connected by bonds with torsional freedomto enable such ligands to twist
and become isohelical with the DNA minor groove. They thus provide optimal shape complementarity with
the DNA receptor and in most cases a typical crescent shape is adopted by these ligands (Figure 9.13).
The DNA minor groove is narrower in A T-rich sequences compared to G C-rich sequences and thus cor-
rectlytwisted aromatic rings in minor groove-binding ligands fit better at into the A T minor groove to
make optimal van der Waals contacts with the helical chains that define the walls of the groove. Additional
specificity (and to some extent stability) is derived from molecular contacts between the bound ligand and
the edges of the base pairs on the floor of the groove. Hydrogen bonds can be accepted by A T base pairs
from the bound molecule to the C-2 carbonyl oxygen of thymineor the N-3 nitrogen adenine. Even though
similar functional groups are available in G C base pairs, the amino group of guanine presents a steric
block to hydrogen bond formation at N-3 of guanine and at the O-2 carbonyl of cytosine. The inter-base
hydrogen bond between the guanine amino group and the cytosine carbonyl oxygen lies in the minor groove
and this interaction sterically inhibits penetration of small molecules into the minor groove at G C sites.
Hence the aromatic rings of many minor groove-binding ligands form close contacts with A-H-2 protons in
the minor groove, but there is no room for the added steric bulk of the guanine amino group in G C base pairs.
Bernard Pullman has shown that negative electrostatic potential is greater in A T minor groovesthan it is
in G C minor grooves. This provides an additional important source for the observed A T specificity of many
minor groove-binding ligands, which often are positively charged. Synthesis of ligands capable of accepting
hydrogen bonds from the guanine amino group is one way of enhancing minor groove-binding at G C sites.
Unlike most intercalators, groove-binding molecules can extend to span many base pairs along the
groove and hence they can exhibit very high levels of DNA sequence-specific recognition. A long estab-
lished goal has been to produce molecules that can specifically recognise DNA sequences long enough to
be unique in a biological context, for example, a eukaryotic regulatory sequence. Minor groove-binding
drugs28–30provide some hope of achieving this goal (Section 9.7.4) and some archetypal minor groove lig-
ands are described below.
9.7.2 Netropsin and Distamycin
Netropsinwas first isolated from Streptomyces netropsis by Finlay in 1951 while Arcamone discovered
distamycin Aas a by-product of Streptomyces distallicusfermentation in 1958. Both drugs are
Reversible Small Molecule–Nucleic Acid Interactions 361