Robert J. Kennedy

Large Circular Dichroism Ellipticities for N-Templated Helical Polypeptides Are Inconsistent with Currently Accepted Helicity Algorithms

An unprecedented, high degree of helicity as judged by CD spectroscopy is observed in N-templated model peptides of the type AcHel-(Ala4 Lys)n Ala2 -NH2 (AcHel-Ala peptide pictured; AcHel is an N-terminal helix-inducing template for polypeptides). These results raise concern over the current methods for determining 100 % helicity.

Context-independent, temperature-dependent helical propensities for amino acid residues.

Assigned from data sets measured in water at 2, 25, and 60 degrees C containing (13)C=O NMR chemical shifts and [theta](222) ellipticities, helical propensities are reported for the 20 genetically coded amino acids, as well as for norvaline and norleucine. These have been introduced by chemical synthesis at central sites within length-optimized, spaced, solubilized Ala(19) hosts. The resulting polyalanine-derived, quantitative propensity sets express for each residue its temperature-dependent but context-independent tendency to forego a coil state and join a preexisting helical conformation. At 2 degrees C their rank ordering is: P << G < H < C, T, N < S < Y, F, W < V, D < K < Q < I < R, M < L < E < A; at 60 degrees C the rank becomes: H, P < G < C < R, K < T, Y, F < N, V < S < Q < W, D < I, M < E < A < L. The DeltaDeltaG values, kcal/mol, relative to alanine, for the cluster T, N, S, Y, F, W, V, D, Q, imply that at 2 degrees C all are strong breakers: DeltaDeltaG(mean) = +0.63 +/- 0.11, but at 60 degrees C their breaking tendencies are dramatically attenuated and converge toward the mean: DeltaDeltaG(mean) = +0.25 +/- 0.07. Accurate modeling of helix-rich proteins found in thermophiles, mesophiles, and organisms that flourish near 0 degrees C thus requires appropriately matched propensity sets. Comparisons are offered between the temperature-dependent propensity assignments of this study and those previously assigned by the Scheraga group; the special problems that attend propensity assignments for charged residues are illustrated by lysine guest data; and comparisons of errors in helicity assignments from shifts and ellipticity data show that the former provide superior precision and accuracy.

Chaotropic Anions Strongly Stabilize Short, N‐Capped Uncharged Peptide Helicies: A New Look at the Perchlorate Effect

Regiospecific binding of perchlorate ions to the N-terminus of short-chained template-substituted polyalanine sequences in water dramatically increases helicity.

An amino acid that controls polypeptide helicity: β-amino alanine, the first strongly stabilizing C-terminal helix stop signal

Abstract β-Amino alanine is shown to stop helix propagation in a polyalanine context while effectively stabilizing the resulting C-terminus. Structural evidence is provided by circular dichroism and t/c ratios from the reporting conformational template Ac-Hel.

Helix-coil energetics for helix formers and breakers reflect context and temperature: mutants of helically robust, guest-sensitive homopeptide hosts.

The natural amino acids are primarily helix breakers at the low assignment temperatures characteristic of many studies, but recent genomic analyses of thermophilic proteins suggest that at high temperatures, some breakers may become strong helix formers. Moreover, the breaker/former inventory has not been previously characterized at the physiologically relevant temperature of 37°C. The versatility of 13CO NMR chemical shifts as helicity reporters allows construction of two mutant peptide series, tailored to expand the range of temperature assignments for helical propensities and derived from the core hosts tL‐Ala9XxxAla9‐tL and tL‐AlaNva4XxxNva4Ala9‐tL, Nva = norvaline. For three limiting guests Xxx, the helix former Nva and the breakers Gly and Pro, we report wXxx[T] assignments at seven temperatures from 2 to 80°C, validating our reasoning and paving the way for assignment of a definitive wXxx[T] data‐base.

C-terminal helix capping propensities in a polyalanine context for amino acids bearing nonpolar aliphatic side chains

Abstract Relative C-capping propensities for nonpolar amino acids and the primary amide, which control helicity for many small peptides, have been determined by a new method. Practical consequences of the observed propensities and their temperature dependences are discussed.