Mechanism of Stabilization of Helical Conformations of Polypeptides by Water Containing Trifluoroethanol

Abstract

For conjugates Ac-Hel 1 -Ala n -OH, n ) 1-6, of the previously characterized reporting, conformational template Ac-Hel 1 , increases in helicity induced by trifluoroethanol (TFE) in water have been related to a simple function of the peptide length n, yielding the helix propagation constant s Ala , which increases from 1.0 to 1.5 for χ TFE ) 0-20 mol %. The per-residue helicity increase is similar to the increase in te state stability induced by TFE in monoamide conjugates Ac-Hel 1 -NHR. Addition of TFE to water significantly increases the rate of interconversion of s-cis/s-trans amide conformers for Ac-Pro-NHMe, consistent with a significant and selective destabilization of the planar resonance-stabilized amide. In dilute aqueous solution TFE increases helicity by selectively destabilizing amide functions that are solvent exposed, with the consequence that compact conformations such as helices that maximize intramolecular amide-amide hydrogen bonding and minimize amide solvent exposure are selectively favored.Medium-sized peptides with an intrinsic tendency to assume helical conformations in water often show a dramatic increase in helicity upon addition of relatively low concentrations of certain alcohols, of which the most efficient appears to be trifluoroethanol (TFE). Since its discovery by Goodman and co-workers, 1 this helix-enhancing effect of TFE has found many applications, most recently for NMR-based characterization of short helices derived from natural proteins. 2 Because TFE extends the range of detectable helicity to include polypeptides that are predominantly unstructured in water, titrations with TFE permit comparisons of relative R-helical propensities between members of peptide series or within regions of the same peptide. 3 Helicity changes resulting from TFE titrations of synthetic peptides have been correlated with amino acid composition, sequence, and helicity-inducing local features. 4,5 Unfortunately, TFE titrations are often difficult to interpret, since the mechanism by which TFE acts to enhance helicity is widely viewed as uncertain or unknown. 2d,3a,c,d A definitive helix enhancement mechanism presumes complete characterization of the effects of TFE on the solvation states of both starting materials and products. Characterization at this level is difficult, since key features of the structure of water itself remain undefined. We address the following practical questions in this report: (1) Which of the two global states is energetically perturbed by TFE? Does TFE selectively raise the energy of the nonhelical random coil state, or lower the energy of the helical state, or both? (2) If the helical state is stabilized, does TFE interact directly with the helix and, if so, with which of its functionalities? (3) If TFE acts by decreasing the stability of the coil state, can the overall stability change be approximated as a sum of effects at each amino acid residue? If it can, does the per-residue effect arise primarily from energy changes involving the amino acid side chains or the a...

Probing the self‐association, intermolecular contacts, and folding propensity of amelogenin

“…[28][29][30][31][32][33] For rP172, CD experiments performed at pH 5.8 in the presence of TFE reveal alcohol-dependent global conformational transitions [ Fig. 1(A)].…”

“…28,29 A direct overlay comparison of the 15 N-1 H HSQC AQ-and 70-rP172 spectra reveals resonant frequency mismatch arising from TFE solvent-exchange-and conformationally induced chemical shift effects. [28][29][30][31][32][33][34][35] Given that monomeric rP172 exists in a globally extended conformational state, 9 it is not surprising that this protein is susceptible to TFE -OH proton exchange [28][29][30] at NH backbone and sidechain sites. As a result, TFE and water proton chemical exchange significantly contribute to the differences in 1 H frequency dispersion [ Fig.…”

“…Over the last decade, the technique of ''solvent engineering'' 28 or the use of cosolvent mixtures of water with alcohols such as 2,2,2-trifluoroethanol (TFE) have been employed to change the environment around proteins and mimic certain conditions or to promote of protein folding and self-assembly with reasonable success. [28][29][30][31][32][33] Thus, we chose this approach to simultaneously achieve several goals that cannot be accomplished by other means with IDPs like amelogenin at the present time. These goals are: promote native-like structure within amelogenin, probe disorder-to-order transformations within the three major regions, and at TFE concentrations > 40% v/v [28][29][30][31][32] induce amelogenin oligomerization in a manner that allows identification of intermolecular contacts between amelogenin molecules.…”

Trifluoroethanol effects on helix propensity and electrostatic interactions in the helical peptide from ribonuclease T1

“…However, earlier studies on the role of electrostatic interactions in the stability of the S-peptide from RNase A have shown that magnitude of the charged group effects is unaltered in the presence of TFE (Nelson & Kallenbach, 1986. A general mechanism by which TFE and other aliphatic alcohols could stabilize helices has been suggested (Conio et al, 1970;Storrs et al, 1992;Cammers-Goodwin et al, 1996). These authors suggest that TFE raises the free energy of the random coil state, because TFE-water mixtures are less able to solvate the amide group of the peptide backbone.…”

Limited proteolysis of ribonuclease A with thermolysin in trifluoroethanol

“…The precise physical mechanism of a-helix induction by TFE is not fully understood and is the subject of intense and recent investigations (Thomas & Dill, 1993;Jasanoff & Fersht, 1994;Shiraki et al, 1994;Cammers-Goodwin et al, 1996). Current views are that TFE favors the formation of the intramolecular hydrogen bonds of the a-helical conformation rather than hydrogen bonds to solvent.…”