Binding site size limitations of imidazole-pyrrole polyamides for recognition in the minor groove of DNA

The discovery that the three ring polyamide Im-Py-Py-Dp containing imidazole (Im) and pyrrole (Py) carboxamides binds the DNA sequence 5'-(A,T)G(A,T)C(A,T)-3' as an antiparallel dimer offers a new model for the design of ligands for specific recognition of sequences in the minor groove containing both G,C and A,T base pairs. In Chapter 2, experiments are described in which the sequential addition of five N- methylpyrrolecarboxamides to the imidazole-pyrrole polyamide Im-Py-Py-Dp affords a series of six homologous polyamides, Im-(Py)_2-7-Dp, that differ in the size of their binding site, apparent first order binding affinity, and sequence specificity. These results demonstrate that DNA sequences up to nine base pairs in length can be specifically recognized by imidazole-pyrrole polyamides containing three to seven rings by 2:1 polyamide-DNA complex formation in the minor groove. Recognition of a nine base pair site defines the new lower limit of the binding site size that can be recognized by polyamides containing exclusively imidazole and pyrrolecarboxamides. The results of this study should provide useful guidelines for the design of new polyamides that bind longer DNA sites with enhanced affinity and specificity.

In Chapter 3 the design and synthesis of the hairpin polyamide Im-Py-Im-Py-γ-Im- Py-Im-Py-Dp is described. Quantitative DNase I footprint titration experiments reveal that Im-Py-Im-Py-γ-Im-Py-Im-Py-Dp binds six base pair 5'-(A,T)GCGC(A,T)-3' sequences with 30-fold higher affinity than the unlinked polyamide Im-Py-Im-Py-Dp. The hairpin polyamide does not discriminate between A•T and T•A at the first and sixth positions of the binding site as three sites 5'-TGCGCT-3', 5'-TGCGCA-3', and 5 'AGCGCT- 3' are bound with similar affinity. However, Im-Py-Im-Py-γ-Im-Py-Im-PyDp is specific for and discriminates between G•C and C•G base pairs in the 5'-GCGC-3' core as evidenced by lower affinities for the mismatched sites 5'-AACGCA-3', 5'- TGCGTT-3', 5'-TGCGGT-3', and 5'-ACCGCT-3'.

In Chapter 4, experiments are described in which a kinetically stable hexa-aza Schiff base La³⁺ complex is covalently attached to a Tat(49-72) peptide which has been shown to bind the HIV-1 TAR RNA sequence. Although these metallo-peptides cleave TAR site-specifically in the hexanucleotide loop to afford products consistent with hydrolysis, a series of control experiments suggests that the observed cleavage is not caused by a sequence-specifically bound Tat(49-72)-La(L)³⁺ peptide.

Sterochemically modified polyamides for recognition in the minor groove of DNA

The design of synthetic molecules that recognize specific sequences of DNA is an ongoing challenge in molecular medicine. Cell-permeable small molecules targeting predetermined DNA sequences offer a potential approach for offsetting the abnormal effects of misregulated gene-expression. Over the past twenty years, Professor Peter B. Dervan has developed a set of pairing rules for the rational design of minor groove binding polyamides containing pyrrole (Py), imidazole (Im), and hydroxypyrrole (Hp). Polyamides have illustrated the capability to permeate cells and inhibit transcription of specific genes in vivo. This provides impetus to identify structural elements that expand the repetoire of polyamide motifs with recognition properties comparable to naturally occurring DNA binding proteins. Through the introduction of chiral amino acids, we have developed chiral polyamides with stereochemically regulated binding characteristics. In addition, chiral substituents have facilitated the development of new polyamide motifs that broaden binding site sizes targetable by this class of ligands.

The Art of Madness

The nineteenth century was not an entirely kind time for the female artist. Coming of age as the 1800’s bridged into its latter half, literary artists Elizabeth Stuart Phelps Ward, Charlotte Perkins Gilman, and Kate Chopin were all well aware of their uncharitable culture. Equipped with firm feminist bents and creative visions, each of three women produced a seminal work – The Story of Avis, “The Yellow Wallpaper,” and The Awakening, respectively – taking that atmosphere to task. In these stories, each of the three women produces a female protagonist who struggles for having been born simultaneously an artist and a woman. The writers pit their women’s desires against the restrictive latitude of their time and show how such conditions drive women to madness, as a result of which they are forced to either escape into the blind mind of insanity or deal daily with their pain and inescapable societal condemnation. In an age where “hysteria” was a frequent hit in the vernacular, Phelps, Gilman and Chopin use art and literature as mediums to show that, indeed, there is a method behind the madness.

Strength of thin-walled elliptical cylinders supported at the minor axis

In this investigation it was found that the instability failure of curved sheet is nearly independent of the type of loading and is primarily a function of the maximum stress, radius-thickness ration and modulus of elasticity. A method of correlating the critical stress of thin sheet under several different types of loading is given. An explanation for the experimental critical stress of thin walled cylinders under bending being greater than that for pure compression is given. The strength of unstiffened thin walled circular nose sections under pure bending was found to be controlled by local instability of the section, rather than a large scale instability. The equation of local instability of curved sheet gives values which are in fair agreement with those found experimentally.

The strength of elliptical cylinders supported at the minor axis under bending plus shear loads is governed primarily by the bending strength, and is little effected by the sheer force unless the amount of shear is quite large with respect to the moment. The effect of increasing the amount of elliptically greatly reduces the bending and shear strength of nose sections. Under torsional loads the stress at buckling falls off as the ration of the major to minor axis increases but the failure stress decreases at a slower rate than the buckling stress. The length effect of semi-circular sections under torsion is similar to that of a circular tube, and can be obtained by Donnell's theoretical equation.