Toward threading polyintercalators with programmed sequence specificity
Abstract
Synthetic molecules that can bind DNA in a sequence specific manner are
an interesting area of molecular recognition research with the potential to develop
new therapeutics. A novel class of polyintercalators, composed of 1,4,5,8-
naphthalene tetracarboxylic diimide (NDI) intercalating units connected via
peptide linkers, have been developed in the Iverson research group. A previous
combinatorial library screen identified two different bis-intercalators with altered
sequence specificity and subsequent structural studies revealed that these
molecules have a unique threading mode of polyintercalation, in which the tethering linker can be in either the major or minor groove, depending on the
peptide linker composition.
The peptide linkers are believed to play an important role in the
recognition of target sequences and several analogues were investigated using
DNase I footprinting experiments. These results showed that the position of a
lysine residue in the linker plays a crucial role in recognizing internal sequences
of 5’-GGNNCC-3’ on DNA. Among those derivatives, the parent compound
(1), H2N-Lys-NDI-Gly-Gly-Gly-Lys-NDI-Gly-OH, showed dramatic sequence
reading ability on 5’-GGNNCC-3’ sequences with the order of GGTACC >
GGCGCC > GGATCC ~ GGGCCC with Kd ~ 125 nM.
The previous structural knowledge of the two different bis-intercalators (1,
and 1.2) served as binding modules that enabled us to design threading
polyintercalators with sequence specific contacts in both grooves. A trimer
(4.1), a hybrid molecule of the two bis-intercalator modules was designed to block
both grooves and its programmed sequence reading ability was proved in
footprinting experiments. As a parallel approach to achieve polyintercalation
with programmed sequences, a symmetric tetramer (5.1) that is a conjugate of two
identical bis-intercalator units (1.2) with flexible aliphatic chain linker was
designed. 1D 1
H-NMR titration experiment revealed that this compound binds
to a 14 bp DNA sequence d(GATAAGTACTTATC)2 with 1:1 stoichiometry.
2D 1
H-NMR studies combined with restrained molecular dynamics confirmed a threading binding mode in the three-dimensional structure of the DNA-ligand
complex in which β-alanine residues in one type of linker reside in the minor
groove as expected, while an adipic acid linker resides in the major groove also as
expected. This represents the first structural analysis of any DNA intercalating
molecules larger than a bis-intercalator and confirms that the sequence specific
binding of longer threading polyintercalators can be predicted from bisintercalator modules. These results will provide the molecular basis for the
DNA binding properties of a new generation of threading polyintercalators.