An incipient 310 helix in Piv-Pro-Pro-Ala-NHMe as a model for peptide folding

The molecular mechanism of helix nucleation in peptides and proteins is not yet understood and the question of whether sharp turns in the polypeptide backbone serve as nuclei for protein folding has evoked controversy1,2. A recent study of the conformation of a tetrapeptide containing the stereochemically constrained residue alpha-aminoisobutyric acid, both in solution and the solid state, yielded a structure consisting of two consecutive beta-turns, leading to an incipient 310 helical conformation3,4. This led us to speculate that specific tri- and tetra-peptide sequences may indeed provide a helical twist to the amino-terminal segment of helical regions in proteins and provide a nucleation site for further propagation. The transformation from a 310 helical structure to an alpha-helix should be facile and requires only small changes in the phi and psi conformational angles and a rearrangement of the hydrogen bonding pattern5. If such a mechanism is involved then it should be possible to isolate an incipient 310 helical conformation in a tripeptide amide or tetrapeptide sequence, based purely on the driving force derived from short-range interactions. We have synthesised and studied the model peptide pivaloyl-Pro-Pro-Ala-NHMe (compound I) and provide here spectroscopic evidence for a 310 helical conformation in compound I

Rotational isomerism about the Ca-CO bond in proline derivatives. 1H and 13C NMR studies of benzyloxycarbonyl-Pro-N-methylamide and pivaloyl-Pro-N-methylamide

The 270 MHz 1H n.m.r. spectrum of benzyloxycarbonyl-Pro-N-methylamide in CDCl3 is exchange broadened at 293° K. Spectral lines due to two species are frozen out at 253° K and a dynamically averaged spectrum is obtained at 323° K. A selective broadening of the Cβ and Cγ resonances in the 13C n.m.r. spectrum is observed at 253° K, with a splitting of the Cβ and Cγ resonances into a pair of lines of unequal intensity. A similar broadening of Cβ and Cγ peaks is also detected in pivaloyl-Pro-N-methylamide where cis-trans interconversion about the imide bond is precluded by the bulky t-butyl group. The rate process is thus attributed to rotation about the Cα-CO bond (ψ) and a barrier (ΔG#) of 14kcal mol-1 is estimated. 13C n.m.r. data for pivaloyl-Pro-N-methylamide in a number of solvents is presented and the differences in the Cβ and Cγ chemical shifts are interpreted in terms of rotational isomerism about the Cα-CO bond

X-Pro Peptides: Solution and Solid-state Conformation of Benzyloxycarbonyl−(Aib−Pro)2−methylEsterBenzyloxycarbonyl-{(Aib-Pro)}_2-methyl Ester, a Type I \beta-Turn

The synthesis of the tetrapeptide $benzyloxycarbonyl{(\alpha-aminoisobutyryl-L-prolyl)}_2-methyl ester$ $(Z-{(Aib-Pro)}_2-OMe)$ and an analysis of its conformation in solution and the solid state are reported. Stepwise synthesis using dicyclohexylcarbodiimide leads to racemization at Pro(2). Evidence for the presence of diastereomeric tetrapeptides is obtained from 270-${MHz}^1H-nmr$ and 67.89-MHz 13C-nmr. The all-L tetrapeptide is obtained by fractional crystallization from ethyl acetate. The NH of Aib(3) is shown to be involved in an intramolecular hydrogen bond by variable-temperature 1H-nmr and the solvent dependence of NH chemical shifts. The results are consistent with a \beta-turn conformation with Aib(1) and Pro(2) at the corners stabilized by a 4 \rightarrow 1 hydrogen bond. The molecule crystallizes in the space group ${P2}_12_12_1$, with a = 8.839, b = 14.938, and c = 22.015 A. The structure has been refined to an R value of 0.051. The peptide backbone is all-trans, and a 4 \rightarrow 1 hydrogen bond, between the CO group of the urethane moiety and Aib(3) NH, is observed. Aib(1) and Pro(2) occupy the corner positions of a type I \beta-turn with \phi = -55.4 deg, \psi = -31.3 deg for Aib(1) and \phi = -71.6 deg, \psi = -38 deg for Pro(2). The tertiary amide unit linking Pro(2) and Aib(3) is significantly distorted from planarity (${\Delta}_{\omega}$ = 14.3 deg)

<i>n</i>→π* interactions in proteins

Hydrogen bonds between backbone amides are common in folded proteins. Here, we show that an intimate interaction between backbone amides likewise arises from the delocalization of a lone pair of electrons (n) from an oxygen atom to the antibonding orbital (π*) of the subsequent carbonyl group. Natural bond orbital analysis predicted significant n→π* interactions in certain regions of the Ramachandran plot. These predictions were validated by a statistical analysis of a large, non-redundant subset of protein structures determined to high resolution. The correlation between these two independent studies is striking. Moreover, the n→π* interactions are abundant, and especially prevalent in common secondary structures such as α-, 3(10)-, and polyproline II helices, and twisted β-sheets. In addition to their evident effects on protein structure and stability, n→π* interactions could play important roles in protein folding and function, and merit inclusion in computational force fields