Enhanced Metabolic Stability and Protein‐Binding Properties of Artificial α Helices Derived from a Hydrogen‐Bond Surrogate: Application to Bcl‐xL

Abstract

Keywordshelical structures; hydrogen bonds; molecular recognition; peptidomimetics; protein-protein interactionsThe α helix plays a fundamental role in imparting specificity to protein-protein and proteinnucleic acid interactions. Molecules that can predictably and selectively disrupt these interactions would be invaluable as tools in molecular biology and, potentially, as leads in drug discovery.[1] We recently described a new strategy for the synthesis of artificial α helices in which one main-chain i to i + 4 hydrogen bond in the target α helix is replaced with a carboncarbon bond derived from a ring-closing metathesis reaction (Figure 1). [2] A key feature of this hydrogen-bond surrogate (HBS) approach is that the internal placement of the cross-link affords short helices with minimal perturbations to their molecular recognition surfaces. This method differs significantly from the commonly employed side-chain cross-linking method for helix stabilization. A limitation of the latter approach is that side-chain functionality must be sacrificed to nucleate stable helical conformations. The modified side chains are unavailable for molecular recognition; moreover, the resulting tether blocks at least one face of the putative helix. The HBS approach uniquely allows the synthesis of artificial helices with all side chains available for molecular recognition, and does not place any steric encumbrances on the helix surface. We believe that our artificial α helices have the potential to target protein receptors and regulate protein-protein interactions more successfully than helices with cross-linked side chains.Our initial studies demonstrated that the HBS approach affords highly stable α helices from alanine-rich peptide sequences. Herein, we show that this metathesis-based method can effectively stabilize α-helical conformations in biologically relevant sequences, that the resulting molecules resist proteolytic degradation as compared with the unconstrained analogues, and that the HBS helices can bind a protein target with high affinity. For these proofof-principle protein-binding studies with our artificial helices, we chose to target the extensively studied α helix binding protein, Bcl-xL.[3] Bcl-xL is an antiapoptotic protein that regulates cell death by binding the α-helical BH3 domain of a family of proapoptotic proteins (including Bak, Bad, Bid, and Bax).[4-6] NMR spectroscopic studies by Fesik and co-workers have shown that the 16-mer peptide 1 derived from the Bak BH3 domain adopts an α-helical conformation upon binding to Several methods that afford stabilized α helices or helix mimetics have already been used to target Bcl-xL, thus allowing us to directly compare the performance of our internally constrained artificial α helices. [8][9][10] Significantly, Huang and co-workers recently reported that Bak BH3 α helices stabilized by a lactam-based side-chain cross-linking strategy were unable to bind Bcl-xl.[11] The authors speculated that the lack of binding might be a result of steric clashes between the c...

Molecular docking studies of anti-apoptotic BCL-2, BCL-XL, and MCL-1 proteins with ginsenosides from Panax ginseng

“…BCL2 family proteins can be divided into two classes, one involved in the promotion (pro-apoptotic) and another involved in the inhibition (anti-apoptotic) of apoptosis. The fine balance between the concentrations of pro-apoptotic and anti-apoptotic BCL-2 proteins is maintained by homeostasis; any perturbations to this balance may lead to disease including restenosis, stroke, heart failure, HIV infection, and cancer 3,4 . The anti-apoptotic proteins such as BCL-2, BCL-XL, and MCL1 contain four conserved BH (BCL 2 homology) homology domains such as BH1, BH2, BH3, and BH4, while pro-apoptotic proteins contain only BH3 domains 2,5 .…”

Designer macrocyclic organo-peptide hybrids inhibit the interaction between p53 and HDM2/X by accommodating a functional α-helix

“…2 Reflecting its abundance in protein structures, α-helices are often encountered at the interface of protein-protein complexes. 3 Accordingly, a number of strategies have been developed for stabilization of α-helical peptides 4 , which include the use of hydrogen bond surrogates 5 as well as of a variety of inter-side-chain linkages such as disulfide, 6 lactam, 7 , thioether 8 or triazole 9 bridges, ‘hydrocarbon staples’ 10 , and cysteine cross-linking moieties. 11 …”

17 The protein epitope mimetic approach to protein-protein interaction inhibitors

“…Some of the approaches used so far to stabilize helical conformations in peptides include the use of intramolecular hydrogen-bond surrogates , such as hydrazone or alkenyl links (Cabezas and Satterthwait 1999;Chapman et al 2004;Henchey et al 2010;Patgiri et al 2008;Vernall et al 2009;Wang et al 2005Wang et al , 2006Wang et al , 2008, and helical end-capping groups (Figure 17.3) (Austin et al 1997;Curran 1988a, 1988b; Kemp et al , 1996Lewis et al 1998;Maison et al 2001;Obrecht et al 1999;Schneider and DeGrado 1998). Other methods to enhance helicity in peptides include incorporating unnatural amino acids (Andrews and Tabor 1999), in particular, α-disubstituted amino acids such as Aib (Venkatraman et al 2001) and other α-alkylated-α-amino acids.…”