α,β hybrid peptides: A polypeptide helix with a central segment containing two consecutive β-amino acid residues

Abstract

Conformational studies on the synthetic 11-aa peptide t-butoxycarbonyl (Boc)-Val-Ala-Phe-␣-aminoisobutyric acid (Aib)-(R)-␤ 3 -homovaline (␤Val)-(S)-␤ 3 -homophenylalanine (␤Phe)-Aib-Val-AlaPhe-Aib-methyl ester (OMe) (peptide 1; ␤Val and ␤Phe are ␤ amino acids generated by homologation of the corresponding L-residues) establish that insertion of two consecutive ␤ residues into a polypeptide helix can be accomplished without significant structural distortion. Crystal-structure analysis reveals a continuous helical conformation encompassing the segment of residues 2-10 of peptide 1. At the site of insertion of the ␤␤ segment, helical hydrogen-bonded rings are expanded. A C15 hydrogen bond for the ␣␤␤ segment and two C14 hydrogen bonds for the ␣␣␤ or ␤␣␣ segments have been characterized. The following conformational angles were determined from the crystal structure for the ␤ residues: ␤Val-5 ( ‫؍‬ ؊126°, ‫؍‬ 76°, and ‫؍‬ ؊124) and ␤Phe-6 ( ‫؍‬ ؊88°, ‫؍‬ 80°, and ‫؍‬ ؊118). The N terminus of the peptide is partially unfolded in crystals. The 500-MHz 1 H-NMR studies establish a continuous helix over the entire length of the peptide in CDCl 3 solution, as evidenced by diagnostic nuclear Overhauser effects. The presence of seven intramolecular hydrogen bonds is also established by using solvent dependence of NH chemical shifts.␣͞␤-helix ͉ C14 hydrogen bond ͉ C15 hydrogen bond ͉ ␣␤␤ segment ͉ ␤␤␣ segment T he rapid advances made in elucidating the conformational properties of ␤ amino acid residues (1-4) permit attempts to rationally design hybrid ␣͞␤ peptides, in which guest residues can be incorporated into regular host secondary structures (5). The ␤ residues have been incorporated into both the turn and strand positions of designed ␤-hairpin peptides. There are few examples of the insertion of ␤ residues into well defined ␣-peptide helices. The only crystallographically characterized examples are the structures of 8-and 11-aa peptides, in which a ␤␥ segment has been inserted into a peptide helix, with concomitant expansion of the hydrogen bonded rings at the site of insertion (6). Regular helical structures with mixed hydrogen bonds have been proposed from the NMR studies of alternating ␣͞␤ sequences, containing the stereochemically restricted ␤ residue trans-2-aminocyclopentanecarboxylic acid (ACPC) (7). As part of a program to insert segments containing multiple ␤ residues into ␣-peptide helices, we obtained the 11-aa peptide t-butoxycarbonyl (Boc)-Val-Ala-Phe-aminoisobutyric acid (Aib)-(R)-␤ 3 -homovaline (␤Val)-␤Phe-Aib-Val-Ala-Phe-Aib-OMe 1. This sequence was based on the parent ␣ peptide Boc-Val-Ala-PheAib-Val-Ala-Phe-Aib-Val-Ala-Phe-Aib-OMe 2, which adopted a complete helical conformation in crystals (8). Peptide 1 differs from the parent all-␣ sequence in having the central Val-AlaPhe-Aib segment replaced by a ␤Val-␤Phe-Aib segment, which formally corresponds to replacing a segment of 12 backbone atoms by a unit containing 11 backbone atoms. In this article, we establish the continuous helical conformation of...

Interplay among side chain sequence, backbone composition, and residue rigidification in polypeptide folding and assembly

“…Past work on the folding behavior of peptides and proteins indicates that limited backbone modification in a given sequence, including replacement of one or two ␣-residues with ␤-residues, can be accomplished without significantly altering native structure or function (11)(12)(13). More recently, we have shown that a particular type of nonnatural backbone, composed of a mixture of ␣-and ␤-amino acid residues in an ␣␣␤␣␣␣␤ pattern, can partially mimic the folding and self-assembly behavior of parent ␣-peptide side-chain sequences (14,15).…”

Differentiation of Positional Isomers of Hybrid Peptides Containing Repeats of β-Nucleoside Derived Amino Acid (β-Nda-) and L-Amino Acids by Positive and Negative Ion Electrospray Ionization Tandem Mass Spectrometry (ESI-MSn)

“…The positive ion ESI mass spectra of all these peptides (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20) + ions (Scheme 2). Fragmentation of these peptides can be explained by using the nomenclature, which was originally proposed for α-peptides by Roepstorff and Fohlman, and later modified by Biemann [18][19][20].…”

“…All the dipeptides were eluted from the column with the mobile phase ratio of 2:98 (CH 3 OH:CHCl 3 ) except for the dipeptides 5 and 6, which were eluted with the mobile phase ratio of 3:7 (hexane:ethyl acetate). The tetra-and hexapeptides were eluted from the column with the mobile phase ratio of 4:96 (CH 3 OH: CHCl 3 ) , 6:94 (CH 3 OH:CHCl 3 ), respectively.…”

“…Oligomers of non-natural peptidic residues [4][5][6], particularly with β-amino acids [7,8] and their hybrids with natural L-amino acids have emerged as versatile structural templates (foldamers) as these exhibit predictable and welldefined secondary structures such as helices, turns, and strands, and offer a variety of possibilities for orienting functional sidechains [9][10][11][12][13]. The oligomers of these amino acids both in cisand trans-conformation have exhibited diverse helical patterns in solution [12][13][14].…”

Sophistication of foldamer form and function in vitro and in vivo

“…This strategy relied on the previously observed tolerance of protein secondary-structure to incorporation of β-amino acids [64][65][66][67], which were substituted into the primary sequence of GCN4-p1 and GCN4-pLI [68] (9 of the 33 residues) at positions distal to the interacting interface to avoid destabilizing the dimer or tetramer interface, respectively. The peptide based on GCN4-p1 associated as a trimer via x-ray crystallography, but was monomeric at concentrations sampled via analytical ultracentrifugation (AUC), whereas the α/β-version of GCN4-pLI associated as a trimer by AUC and a tetramer by x-ray crystallography (Figures 3e and f) [57•].…”