Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2006 Jun 28;128(25):8227-33.
doi: 10.1021/ja060094y.

Temperature- and length-dependent energetics of formation for polyalanine helices in water: assignment of w(Ala)(n,T) and temperature-dependent CD ellipticity standards

Affiliations

Temperature- and length-dependent energetics of formation for polyalanine helices in water: assignment of w(Ala)(n,T) and temperature-dependent CD ellipticity standards

Gabriel E Job et al. J Am Chem Soc. .

Abstract

Length-dependent helical propensities w(Ala)(n,T) at T = 10, 25, and 60 degrees C are assigned from t/c values and NMR 13C chemical shifts for series 1 peptides TrpLys(m)Inp2(t)Leu-Ala(n)(t)LeuInp2Lys(m)NH2, n = 15, 19, and 25, m = 5, in water. Van't Hoff analysis of w(Ala)(n,T) show that alpha-helix formation is primarily enthalpy-driven. For series 2 peptides Ac-Trp Lys5Inp2(t)Leu-(beta)AspHel-Ala(n)-beta-(t)LeuInp2Lys5NH2, n = 12 and 22, which contain exceptionally helical Ala(n) cores, protection factor-derived fractional helicities FH are assigned in the range 10-30 degrees C in water and used to calibrate temperature-dependent CD ellipticities [theta](lambda,H,n,T). These are applied to CD data for series 1 peptides, 12 < or = n < or = 45, to confirm the w(Ala)(n,T) assignments at T = 25 and 60 degrees C. The [theta](lambda,H,n,T) are temperature dependent within the wavelength region, 222 +/- 12 nm, and yield a temperature correction for calculation of FH from experimental values of [theta](222,n,T,Exp).

PubMed Disclaimer

Figures

Figure 1
Figure 1
Red dots correspond to t/c data for the series Ac–Hel–AlantLInp2–Lys4Trp–NH2 n = 4–14, in water at 25 °C. Lines plot t/c values L–R-modeled from recursively assigned wAla(n,25) values. Previous parameter values at 2 °C were used to model the upper blue line: A = 0.80; B = 0.17, the initiation parameter for the t-state. The lower blue line and the best-fit green correspond to B = 0.15 and 0.16; j = 6.5, the N-capping Hel parameter for cs-state helices.
Figure 2
Figure 2
Temperature-dependent helical propensities wAla(n,T) for alanine residues in water within polyalanine contexts 1a,b; blue: T = 2 °C; cyan: T = 10 °C; green: T = 25 °C; red: T = 60 °C. Values assigned at or below 25 °C show a positive length dependence, but values assigned at 60 °C show a negative length dependence. CD data (shown below) show that for n > 25, all wAla(n,T) converge to an upper limit.
Figure 3
Figure 3
(a) Experimental per-residue molar ellipticities [θ]λ,22,T,Exp, units, for the peptide Ac–Hel(Ala4Lys)4Ala2NH2 in 32 mol % ethylene glycol–water at intervals of 10 °C from −20 °C (top green curve) to −50 °C (bottom blue curve). Convergence to temperature-independent values of [θ]λ is observed, except within the λ region: 222 ± 12 nm; the slope at 222 nm is +260 deg cm2 K−1 dmol−1. (b) Calculated values of [θ]λ,H,n,T for the completely helical conformations of the series 2 peptide βAspHel–Ala22beta in water at 2 (blue curve), 10 (green curve), and 30 °C (red curve). Units: 103 deg cm2 dmol−1.
Figure 4
Figure 4
Temperature dependences of Figure 3b per-residue molar ellipticities [θ]222,H,22,T (blue) and [θ]208,H,22,T (green) for series 2 Ala22. Linear temperature regressions for Ala22: at 222 nm: intercept −49.7, slope +260; at 208 nm: intercept −34.6, slope +110. Linear regressions for Ala12 (data not shown): at 222 nm, intercept −44.5, slope +280; at 208 nm, intercept −34.7, slope +92. Units: intercept, 103 deg cm2 dmol−1; slope, deg cm2 dmol−1 K−1. Error limits are calculated from errors in FH values.
Figure 5
Figure 5
CD tests of wAla(n,T) data sets using [θ]222 data (green, 25 °C; red, 60 °C) measured in water for Alan 1a peptides.6 L–R modeling of [θ]222 at 25 °C (green line) used [θ]222,n,25 = (−60 500 + 260 (25−2))(1 − X/n), X = 6.5, as assigned at 2 °C. (fit: n = 18–45, SD: 1.2). At 60 °C (red line), [θ]222,n,60 = (−60 500 + 260 (60−2))(1 − X/n), X = 2.5. (fit: n = 16–45, SD: 0.5). Alternative (not shown): [θ]222,n,60 = (−60 500 + 260 (25−2))(1 − X/n), X = 3.7. (n = 16–45, SD: 0.9). Units: 103 deg cm2 dmol−1.
Figure 6
Figure 6
(a) A dual-wavelength parametric plot of cap-corrected [θ]222 vs [θ]208 measured in water for the series 1a peptide. Temperature is the implicit variable, and its range is 60–2 °C (respectively low and high −[θ] values), at intervals of 5 °C from 60 to 5 °C. Three local slopes are shown; 1.5 red; 1.8 green; 2.5, blue; each defined by three or four successive data points at high, medium, and low temperatures. The continuously increasing local slopes reflect the heterogeneity of helical lengths within the conformational ensemble. (The Ala28 spectra exhibit a 203 nm isoelliptic point.6) (b) Series 2 peptide βAspHel–Ala22beta plot exhibits two regions of constant slope: 60–35 °C (red line and data points; C. C. = 0.997; slope 1.8) and 30–2 °C (blue line and data points; C. C. = 0.996; slope 3.2). Units: 103 deg cm2 dmol−1.
Figure 7
Figure 7
Parameter dependences of Lifson–Roig-modeled fractional helicities FH for Ala22 and Ala12 peptides. The temperature dependence of FH values is modeled by varying the length-independent values for w. Magenta lines are calculated for Ala22, orange lines for Ala12. Continuous lines correspond to FH for strong cap stabilization: j = 200, c = 7; broken lines correspond to weak cap destabilization: j = c = 0.5. Symbols drawn as squares (Ala22) and circles (Ala12) report FH calculated from the wAla(n,T) of Figure 2: red: T = 60 °C, green: T = 25 °C, blue: T = 2 °C. Filled symbols correspond to j = 200, c = 7; open symbols correspond to j = c = 0.5. Symbols have been positioned on appropriate line sites at which FH values are equal; thus, an average w (and temperature) is assigned to the wAla(n,T)-derived FH. These sites thus define approximate fixed-w assignments for the four peptides.
None

References

    1. Kennedy RJ, Walker SM, Kemp DS. J Am Chem Soc. 2005;127:16961–16968. (a) - PMC - PubMed
    2. (b) Kennedy. R. J.; Miller, J. S.; Kemp. D. S. Submitted for publication
    1. The fractional helicity FH of a potentially helical peptide is the atom fraction of its backbone α-carbons that belong to helical conformations under particular experimental conditions. The site helicity FHi is the corresponding atom fraction of the α-carbons at a particular residue site i. The average of all FHi over a particular helical region equals the overall FH for that region.
    1. Kallenbach NR, Spek EJ. Methods Enzymol. 1998;285:26–41. For a review through 1997 see: - PubMed
    1. For example: (a) Scheraga, H. A. The Intrinsic Tendency Toward α-Helix Formation. In Perspectives in Structural Biology; Vijayan, M., Yathindra, N., Kolaskar, A. S., Eds.; Indian Academy of Sciences: Bangalore, 1999; pp 275–282.
    2. Creamer TP, Rose GD. Proteins: Struct, Funct, Genet. 1994;19:85–97. (b) - PubMed
    3. Myers JK, Pace CN, Scholtz JM. Biochemistry. 1997;36:10923–10929. (c) - PubMed
    1. Moreau, R. J.; Nasr, K. A.; Török, M.; Miller, J. S.; Schubert, C.; Kennedy, R. J.; Kemp, D. S. In preparation

Publication types