Helix denaturation

Addition of Mg2+ stabilizes polynucleotides100 and the DNA double helix. Denaturation of DNA is accompanied by release of Mg2+. In both cases Mg2+ reduces the electrostatic charge associated with the phosphate groups. 

The value of AH can also be compared to the helix unfolding AH0 of Scholtz et al. (1991). The buried surface area, relative to the extended chain, was calculated for a 50-residue alanine a helix. An average of 19.5 A2 of polar surface is buried per residue and an average of 3.2 A2 of apolar surface is overexposed (i.e., is less accessible in the extended chain than in the helix). Using the fundamental parameters for the polar and apolar ACP described above, a value of — 6.5 cal K-1 (mol res)-1 is estimated for ACp for the helix denatur-ation. At 100°C the extrapolated value of AH0 is about 1.0 kcal (mol res)-1, again in reasonable agreement with the value of AH of 1.35 kcal (mol res)-1. These results strongly support the assertion that the apolar contribution to AH0 is close to zero at 7h. 

Daggett V, Levitt M. Molecular dynamics simulations of helix denaturation. J Mol Biol 1992 223 1121-1138. 

Hypochromism - absorption of light by bases reduced when in helix = denaturation causes increase in absorbance of light at 260 nm. 

Hydrogen bonding is frequently involved in intercalation and interaction of many drugs with the DNA double helix. Denaturation and separation of the component strands will involve rupture of all the inter-base N-H-0 bonding. 

V. Daggett and M. Levitt,/. Mol. Biol., 223,1121 (1992). Molecular Dynamics Simulations of Helix Denaturation. 

FIGURE 9.6 DSC of (a) recombinant resilin in water showing no enthalpic events, (b) bovine serum albumin in phosphate-buffered saline (PBS) showing denaturing occurring at 62°C, and (c) wool fiber in water showing denaturing of the a-helix at 145°C (Endotherm up). 

Fig. 1. Schematic diagram of nuclease A131A in the folded conformation. The alpha helices and beta strands are labeled. NMR analysis suggests the two turns and one helix in black are modestly populated in the denatured state, whereas the shaded helix is slightly populated. Strands / l-/ 2-/ 3 form an extended structure about which littie is known. Reproduced from Barron, L. D., Hecht, L., Blanch, E. W., and Bell, A. F. (2000). Prog. Biophys. Mol Chem. 73, 1-49. 2000, with permission from Elsevier Science.

Fig. 6. Spectral monitoring of the thermal denaturation of the highly helical, Ala-rich peptide Ac-(AAAAK)3AAAA-YNH2 in D20 from 5 to 60°C, as followed by changes in the amide V IR (left) and VCD (right). IR show a clear shift to higher wavenumber from the dominant a-helical peak (here at an unusually low value, 1637 cm-1, due to full solvation of the helix) to a typical random coil value ( 1645 cm-1). VCD loses the (—,+,—) low-temperature helical pattern to yield a broad negative couplet, characteristic of a disordered coil, at high temperature. Spectra were normalized to A = 1.0 by 45°C.

Fig. 12. Thermal denaturation for ribonuclease Tj as followed by VCD, from 20° to 65°C. The matrix descriptors determined for the native state and the unfolded high-temperature data are indicated. The values indicate a loss of the helix segment but maintenance of sheet segments. Also listed are the spectrally determined fractional contributions (FC) to the secondary structure. When combined with the segment analysis, this implies that the residual sheet segments must be very short. Reprinted with permission from Pancoska, P., et al. (1996). Biochemistry 35(40), 13094-13106, the American Chemical Society.

Table V shows the results of this analysis for the Pn-helix fraction of several proteins denatured by heat, cold, acid, and Gdm HCl/urea. There is rather good consistency among the estimated Pn-helix contents for proteins denatured by a given agent, except for acid-denatured proteins, which show more variability. The chemically denatured proteins have 30 5% Pn-helix content near 0°C. At the other extreme, heat-denatured proteins have Pn-helix contents near 0%, with lysozyme having the highest value (8%). Although there are only two examples of cold-denatured proteins in Table V,2 they both have Pn-helix contents of about 20%. Acid-denatured proteins have Pn-helix contents ranging from 0 to 16%.

Proteins unfolded by GdmHCl or urea will have a dominant conformation, Pn- At low temperatures we find about one-third of the residues in chemically denatured proteins in the Pn-helix conformation, with two-thirds in the form of the high-temperature ensemble. Since at least one-third of the residues in this ensemble are isolated Pn residues or in Pn helices of two or three residues, the total Pn content will be 50% or greater. The Pn content of cold- and acid-denatured proteins will be substantial, probably >40%, but not as large as in chemically denatured proteins. 

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