A R COO 15b R Pr

Scheme 3.8

showed significant upfield shifts of aromatic protons, NOE signals between protons on adjacent aromatic units, and diastereotopic methylene hydrogens possibly resulted from restricted rotation due to intramolecular association [99].

Aedamer 15b was synthesized to mimic leucine zippers found in peptide systems [101]. However, aromatic interactions were complicated by the presence of other hydrophobic units (i.e. leucine). Instead of adopting the desired leucine zipper, 16 folded as typical aedamers at low temperatures. Upon heating to 80 °C, an irreversible conformational change happened as a result of intermolecular association. The red wine colored solution turned into a pale, gel-like solution, apparently due to loss of aromatic stacking. With the visible color change, 15b could be potentially useful as a thermal sensor indicating onset of a threshold temperature.

In another study, the authors undertook an extensive investigation in the effect of tethering spacers on the conformation of dimeric model systems [102]. Folded structures were found to dominate with both rigid and flexible spacers varying in length, reflecting the robustness of intramolecular aromatic interactions in aqueous solutions. On the other hand, since the spacer could alter the orientation of the donor and acceptor units, it could be used to deliberately put the aromatic rings out of contact of each other. The result was that aromatic units had to aggregate intermolecularly. For example, the charge-transfer (CT) band and hypo-chromism of 16 decreased dramatically at high temperatures (80 °C), in contrast to all the other folded dimers. Another conclusion from this study was that the stacked conformations of aedamers were highly degenerate, reasonable for molecules with fairly flexible spacers.

With binding most preferably taking place between donors and acceptors, aromatic associations could be exploited to create double-stranded foldamers from an acceptor strand and a donor strand (see Fig. 3.4.19 in Chapter 4) [103]. (Strictly speaking, double-stranded foldamers are stabilized by nonadjacent units. These examples are included together with other aedamers for the convenience of discussion.) Despite strong repulsions between the two negatively charged chains, Ka increased steadily from 1.3 x 102 for strands containing only a single aromatic unit to 3.5 x 105 for those with four. A high degree of discrimination existed in the binding process, as an excess of either the donor or the acceptor strand (n = 4) migrated separately from the complex during polyacrylamide gel electrophoresis.

In addition to forming a duplex with a donor strand, the acceptor strand could do so with a DNA strand. In aedamers 17-19, several lysine residues were introduced to favor electrostatic interactions between the aedamer and DNA. Compounds 17 and 18 were identified from a library of dimeric derivatives to bind DNA with interesting specificity [104]. The former intercalated DNA with the -Gly3-Lys- linker in the major groove [105], while the latter with the -Ala3-Lys-linker in the minor groove [106]. On the basis of these earlier findings, tetramer 19 was designed to bind sequentially in the minor groove, major groove, minor groove, in a manner similar to how a snake might try to climb a ladder (Fig. 3.4). The binding pattern was confirmed by titration studies and NMR spectroscopy [107]. In the future, these molecules may offer significant opportunities in binding long strands of DNA with sequence specificity.

Fig. 3.4 Threading tetraintercalator that binds DNA. (Reprinted with permission from Ref 107. Copyright 2004, American Chemical Society, Washington, DC.)
Scheme 3.10

The donor-acceptor motif was also used by Ramakrishnan et al. in other systems [108, 109]. Polymer 20 (Mn = 30 000-50 000, PDI a 2) had much longer spacers in between the aromatic units compared to those in most other foldamers discussed so far. Polymers with shorter spacers (x = 3) folded better than those with longer ones (x = 4 and 5). Folding of the latter (20b and 20c), however, could be facilitated by the addition of methanol to enhance solvophobic interactions and van der Waals interactions, or with alkali metal ions to contract the oligo(ethylene glycol) tethers.

92 | 3 Foldamers Based on Solvophobic Effects 3.4.3

Foldamers Stabilized by Nonadjacent Aromatic Units

In foldamers 7-11, stacked conformations are either predetermined by the conformations of the linkages or highly favored by the short spacers between the aromatic units. They are unlikely to ''unfold'' under reasonable conditions. This is in direct contrast to most biofoldamers that are characterized by dynamic confor-mational behavior. To achieve folding-unfolding reversibility, one must introduce some flexibility in the structure, most likely at the spacers. This was the case in aedamers, which could be ''denatured'' by cationic surfactant cetyltrimethylam-monium bromide (CTAB) [99]. Nevertheless, the folding motif until now is limited to columns of stacked aromatic rings. With flexible spacers, it is difficult to imagine any folding motifs other than the stacked columns illustrated in Fig. 3.2a, whether identical or different aromatic units are involved. In order to obtain foldamers stabilized by nonadjacent units, one must constrain the chain in a way to avoid association of neighboring units.

Such a strategy was successfully employed by Moore and co-workers in the m-phenylene ethynylene (mPE) foldamers (22) [110]. These foldamers were inspired by their discovery that macrocycle 21 self-associated by face-to-face p-stacking interactions [111]. Of the features important to the design, the most critical was the utilization of a semirigid aromatic backbone. As mentioned previously, semirigid-ity is a strategy universally adopted by nature to simplify the conformational problem. With a 120° angle created by the meta-connectivity and linear ethynylene spacers, an mPE oligomer was geometrically poised to fold upon itself, forming a conformer resembling macrocycle 21 (Fig. 3.5). Tri(ethylene glycol) (Tg) was

Fig. 3.5 Relationship between intermodular aggregation of mPE macrocyles and the intramolecular folding of linear mPE oligomers. (Reprinted with permission from Ref 112. Copyright 2006, American Chemical Society, Washington, DC.)

Fig. 3.5 Relationship between intermodular aggregation of mPE macrocyles and the intramolecular folding of linear mPE oligomers. (Reprinted with permission from Ref 112. Copyright 2006, American Chemical Society, Washington, DC.)

chosen as the side chain because of its good solubility in polar solvents promoting aromatic interactions. Placement of the side chains at the periphery of the envisaged helix creates a hydrophilic shell around the hydrophobic core, a feature universal in water-soluble proteins.

Unlike a-helices found in proteins, synthetic helices do not have known spectroscopic signatures to allow their quick characterization. Since six units were expected to make one turn in 22, a natural anticipation was that a critical chain length should exist. For example, in order to benefit from any intramolecular p-stacking interactions, the chain should be longer than the hexamer. Indeed, under dilute concentrations (e.g. 10 mM) at which intermolecular aggregation was minimized, significant upfield shifts of proton signals occurred abruptly in acetonitrile for 22n with n > 8. In chloroform, a solvent that weakened aromatic interactions, the proton signals were essentially independent of chain lengths [110].

This kind of chain-length dependence test (CLDT) turned out extremely useful for the characterization of mPE foldamers [112]. When the percentage of acetoni-trile was gradually increased in a mixture of acetonitrile and chloroform, absorption for 23n at 289 nm increased while that at 303 nm decreased (Fig. 3.6a, left). These changes were attributed to a shift of the equilibrium toward the cisoid con-formers [113]. A plot of the ratio of absorbances (A303/A289) indicated a distinct transition in acetonitrile with n > 8 (Fig. 3.6a, right). Consistent with the folded, stacked conformation, a broad, excimer-like emission band shifted to the red in acetonitrile and replaced the sharp emission at about 350 nm for oligomers longer than the 10-mer (Fig. 3.6b). Such a chain-length-dependency again was absent in chloroform. When the side chains were made chiral, induced circular dichroism (CD) signals were observed in acetonitrile for 24n with n > 8 (Fig. 3.6c), consistent with helical conformations that were biased in handedness by the asymmetric side chains [114]. During solvent titration experiments, these spectro-scopic changes were found to display a sigmoidal relationship with the solvent composition, reminiscent of denaturation curves used to determine thermo-dynamic stabilities of proteins. The data fit well to a two-state model, in agreement with the proposed helix-coil transition [113]. Spin-labeling experiments later confirmed that six monomers made up one turn in the mPE foldamers [115].

Quite surprisingly, the folded state of the mPE foldamers was found to be stable in a range of polar and nonpolar solvents including ethyl acetate, DMSO, acetonitrile, methanol, TFE, and even reasonably well in nonpolar, polarizable solvents such as carbon tetrachloride [116]. The only solvents that promoted unfolding of the helices were chlorinated solvents such as chloroform, methylene chloride, and 1,2-dichloroethane. When the Tg side chains were replaced with nonpolar alkyl groups, the resulting foldamers folded well in heptane [117]. Apparently, strong intramolecular interactions were present in the mPE foldamers. It is unclear whether van der Waals or solvophobic interactions play the dominant role in the unusual stability of the helix. Solvophobic contributions may be quite substantial, as mPE derivatives in general have very poor solubility in most sol

94 I 3 Foldamers Based on Solvophobic Effects

Fig. 3.6 (a) UV spectra of 2318 (left) and the ratio of absorbances at 303 and 289 nm for 234-23n8 (right). (b) Fluorescence spectra of 23n8 (left) and the ratio of fluorescence intensities at 410 and 350 nm for 234-23ns (right). (c) CD spectra of 24ns (right) and anisotropy factor (Ae/e) at 315 nm for 244-

Fig. 3.6 (a) UV spectra of 2318 (left) and the ratio of absorbances at 303 and 289 nm for 234-23n8 (right). (b) Fluorescence spectra of 23n8 (left) and the ratio of fluorescence intensities at 410 and 350 nm for 234-23ns (right). (c) CD spectra of 24ns (right) and anisotropy factor (Ae/e) at 315 nm for 244-

2418 (right). Data collected in CHCl3 are indicated by blue squares, and those collected in CH3CN are indicated by red circles. (Reprinted with permission from Ref 112. Copyright 2006, American Chemical Society, Washington, DC.)

vents unless bulky or flexible groups are attached. As mentioned before, although partly due to strong solute-solute interactions, low solubility may also result from poor solvation of a rigid framework that does not gain as much entropy as a flexible one during dissolution.

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