Circular dichroism spectra of short, fixed-nucleus alanine helices

Abstract

Very short alanine peptide helices can be studied in a fixed-nucleus, helix-forming system [Siedlicka, M., Goch, G., Ejchart, A., Sticht, H. & Bierzynski, A. (1999) Proc. Natl. Acad. Sci. USA 96, 903-908]. In a 12-residue sequence taken from an EF-hand protein, the four C-terminal peptide units become helical when the peptide binds La 3؉ , and somewhat longer helices may be made by adding alanine residues at the C terminus. The helices studied here contain 4, 8, or 11 peptide units. Surprisingly, these short fixed-nucleus helices remain almost fully helical from 4 to 65°C, according to circular dichroism results reported here, and in agreement with titration calorimetry results reported recently. These peptides are used here to define the circular dichroism properties of short helices, which are needed for accurate measurement of helix propensities. Two striking properties are: (i) the temperature coefficient of mean peptide ellipticity depends strongly on helix length; and (ii) the intensity of the signal decreases much less rapidly with helix length, for very short helices, than supposed in the past. The circular dichroism spectra of the short helices are compared with new theoretical calculations, based on the experimentally determined direction of the NV 1 transition moment.A long-standing goal in the study of peptide helices has been to relate the properties of helix formation in standard peptides (for example, in alanine-based peptides; refs. 1-3) to those observed when the helix is initiated by a helical template or an actual fixed helical nucleus (4-7). The first results obtained by using a synthetic template to initiate helix formation (4-6) showed little relation to the results found with standard peptides. In particular, alanine residues added to the synthetic template were reported (4-6) to have a lower helix propensity than in alanine-based peptides. The synthetic template used in these initial studies was acetyl-L-Pro-L-Pro, only one of whose three conformers is productive in initiating a helix (4-6). Bierzynski and coworkers (7) succeeded in nucleating short alanine-rich helices with a helical nucleus obtained by binding La 3ϩ to a 12-residue peptide (P1) modeled on an EF-hand protein. The four C-terminal peptide units become helical when P1 binds La 3ϩ , and longer helices can be made by adding residues at the C terminus of P1. The NMR structure (7) of a second peptide P2, with four additional residues (A 3 Q) added to, P1 defines the structure of the fixed helical nucleus, and agrees with earlier work (8) on the ligands that coordinate La 3ϩ . All but one of the participating ligands lie outside the helix in the N-terminal direction; the side-chain -COOH group of Glu 12, inside the helix, participates in binding La 3ϩ . Recently, Bierzynski and coworkers made an extensive study of helix propagation in this system (to be published), and they found a helix propensity for alanine in good agreement with results from standard alaninebased peptides (A. Bierzynski, personal communication).Circular dich...

Backbone-methylated Analogues of the Principle Receptor Binding Region of Human Parathyroid Hormone

“…This model resulted in predicted ellipticities of zero for Val 21 to Lys 26 , contrary to the observed presence of the helix after these residues were N-methylated. The more recent model of Chin et al (34), using a dependence of x on helix length (34), resulted in a profile much more similar to the experimentally observed one. This suggests that, even in the presence of backbone N-methylation, short helical regions are present in the 12-residue-long helix in hPTH(1-31)NH 2 .…”

“…Peptide concentrations were calculated from the absorption at 280 nm, using an extinction coefficient of 5700 M Ϫ1 for the single tryptophan. (34) formulation, the x is effectively dependent on N p and results in ellipticities for helical regions with as little as two residues.…”

Influences of temperature and threshold effect of NaCl concentration on Alpias vulpinus OCT

“…Also in the case of the slope of the plot of G* versus NaCl concentrations there is a linear correlation between these two parameters. The slope of the plots depends to the number of sites buried, upon folding, that are available, following unfolding, for the interaction with solvent [31]. Data confirm that there is a tight correlation between exposure of buried sites and NaCl concentration in spite of temperature increase.…”

Enthalpy of helix–coil transition: Missing link in rationalizing the thermodynamics of helix-forming propensities of the amino acid residues

“…The helical fraction in P2X peptides in the absence of La 3ϩ was estimated from the experimentally measured ellipticities of the peptides by using a procedure described before (26,27). As expected, the measured values of ellipticities vary for different peptides, indicating different amounts of the fractional helical structure in P2X (X ϭ A, G, V, L, I, J, B, N, Q, S, T) series of peptides.…”

“…These changes in the structure induced by La 3ϩ binding can also be monitored by CD spectroscopy (26,27). For example, upon La 3ϩ binding, the ellipticity of the P2A peptide changes from Ϫ4,100 to Ϫ11,600 deg⅐cm 2 ⅐dmol Ϫ1 at 25°C.…”