Residue-Based Preorganization of BH3-Derived α/ß-Peptides: Modulating Affinity, Selectivity and Proteolytic Susceptibility in α-Helix Mimics.

We report progress toward a general strategy for mimicking the recognition properties of specific α-helices within natural proteins through the use of oligomers that are less susceptible than conventional peptides to proteolysis. The oligomers contain both α- and ß-amino acid residues, with the density of the ß subunits low enough that an α-helix-like conformation can be adopted but high enough to interfere with protease activity. Previous studies with a different protein-recognition system that suggested ring-constrained ß residues can be superior to flexible ß residues in terms of maximizing α/ß-peptide affinity for a targeted protein surface. Here, we use mimicry of the 18-residue Bim BH3 domain to expand the scope of this strategy. Two significant advances have been achieved. First, we have developed and validated a new ring-constrained ß residue that bears an acidic side chain, which complements previously known analogues that are either hydrophobic or basic. Second, we have discovered that placing cyclic ß residues at sites that make direct contact with partner proteins can lead to substantial discrimination between structurally homologous binding partners, the proteins Bcl-xL and Mcl-1. Overall, this study helps to establish that α/ß-peptides containing ring-preorganized ß residues can reliably provide proteolytically resistant ligands for proteins that naturally evolved to recognize α-helical partners.

Extending foldamer design beyond α-helix mimicry: α/ß-peptide inhibitors of vascular endothelial growth factor signaling.

Diverse strategies have been explored to mimic the surface displayed by an α-helical segment of a protein, with the goal of creating inhibitors of helix-mediated protein-protein interactions. Many recognition surfaces on proteins, however, are topologically more complex and less regular than a single α-helix. We describe efforts to develop peptidic foldamers that bind to the irregular receptor-recognition surface of vascular endothelial growth factor (VEGF). Our approach begins with a 19-residue α-peptide previously reported by Fairbrother et al. (Biochemistry 1998, 37, 17754) to bind to this surface on VEGF. Systematic evaluation of αâ†’ß replacements throughout this 19-mer sequence enabled us to identify homologues that contain up to ~30% ß residues, retain significant affinity for VEGF, and display substantial resistance to proteolysis. These α/ß-peptides can block VEGF-stimulated proliferation of human umbilical vein endothelial cells.

Evaluation of diverse α/ß-backbone patterns for functional α-helix mimicry: analogues of the Bim BH3 domain.

Peptidic oligomers that contain both α- and ß-amino acid residues, in regular patterns throughout the backbone, are emerging as structural mimics of α-helix-forming conventional peptides (composed exclusively of α-amino acid residues). Here we describe a comprehensive evaluation of diverse α/ß-peptide homologues of the Bim BH3 domain in terms of their ability to bind to the BH3-recognition sites on two partner proteins, Bcl-x(L) and Mcl-1. These proteins are members of the anti-apoptotic Bcl-2 family, and both bind tightly to the Bim BH3 domain itself. All α/ß-peptide homologues retain the side-chain sequence of the Bim BH3 domain, but each homologue contains periodic α-residue → ß(3)-residue substitutions. Previous work has shown that the ααßαααß pattern, which aligns the ß(3)-residues in a 'stripe' along one side of the helix, can support functional α-helix mimicry, and the results reported here strengthen this conclusion. The present study provides the first evaluation of functional mimicry by ααß and αααß patterns, which cause the ß(3)-residues to spiral around the helix periphery. We find that the αααß pattern can support effective mimicry of the Bim BH3 domain, as manifested by the crystal structure of an α/ß-peptide bound to Bcl-x(L), affinity for a variety of Bcl-2 family proteins, and induction of apoptotic signaling in mouse embryonic fibroblast extracts. The best αααß homologue shows substantial protection from proteolytic degradation relative to the Bim BH3 α-peptide.

A partner evokes latent differences between Hox proteins.

Hox transcription factors bind highly related DNA sequences in vitro, yet they regulate different genes and play distinct roles in anterior-posterior patterning in animals. Slattery et al. report that a common cofactor, Exd, accentuates latent sequence specificities of all eight Hox proteins and directs binding to relevant sites across the genome.

Small-molecule regulators that mimic transcription factors.

Transcription factors (TFs) are responsible for decoding and expressing the information stored in the genome, which dictates cellular function. Creating artificial transcription factors (ATFs) that mimic endogenous TFs is a major goal at the interface of biology, chemistry, and molecular medicine. Such molecular tools will be essential for deciphering and manipulating transcriptional networks that lead to particular cellular states. In this minireview, the framework for the design of functional ATFs is presented and current challenges in the successful implementation of ATFs are discussed.

Nucleating the assembly of macromolecular complexes.

Nature constructs intricate complexes containing numerous binding partners in order to direct a variety of cellular processes. Researchers have taken a cue from these events to develop synthetic molecules that can nucleate natural and unnatural interactions for a diverse set of applications. These molecules can be designed to drive protein dimerization or to modulate the interactions between proteins, lipids, DNA, or RNA and thereby alter cellular pathways. A variety of components within the cellular machinery can be recruited with or replaced by synthetic compounds. Directing the formation of multicomponent complexes with new synthetic molecules can allow unprecedented control over the cellular machinery.

High-resolution structural characterization of a helical alpha/beta-peptide foldamer bound to the anti-apoptotic protein Bcl-xL.

Get into the groove: The first high-resolution structure of a foldamer bound to a protein target is described (see picture; foldamer in sticks). The foldamer consists of alpha- and beta-amino acid residues and is bound to the anti-apoptotic protein Bcl-x(L). The overall binding mode and key interactions observed in the foldamer/Bcl-x(L) complex mimic those seen in complexes of Bcl-x(L) with natural alpha-peptide ligands. Additional contacts in the foldamer/Bcl-x(L) complex involving beta-amino acid residues appear to contribute to binding affinity.