Random-coil behavior and the dimensions of chemically unfolded proteins

Abstract

Spectroscopic studies have identified a number of proteins that appear to retain significant residual structure under even strongly denaturing conditions. Intrinsic viscosity, hydrodynamic radii, and small-angle x-ray scattering studies, in contrast, indicate that the dimensions of most chemically denatured proteins scale with polypeptide length by means of the power-law relationship expected for random-coil behavior. Here we further explore this discrepancy by expanding the length range of characterized denatured-state radii of gyration (R(G)) and by reexamining proteins that reportedly do not fit the expected dimensional scaling. We find that only 2 of 28 crosslink-free, prosthetic-group-free, chemically denatured polypeptides deviate significantly from a power-law relationship with polymer length. The R(G) of the remaining 26 polypeptides, which range from 16 to 549 residues, are well fitted (r(2) = 0.988) by a power-law relationship with a best-fit exponent, 0.598 +/- 0.028, coinciding closely with the 0.588 predicted for an excluded volume random coil. Therefore, it appears that the mean dimensions of the large majority of chemically denatured proteins are effectively indistinguishable from the mean dimensions of a random-coil ensemble.

Fig. 1.

The R_G of the large majority of chemically denatured proteins scale with polymer length, N, by means of the power-law relationship R_G = R₀N^ν. Two statistically significant outliers, creatine kinase and angiotensin II, are indicated. The solid line, which is the least-squares fit ignoring the two potential outliers, produces an exponent, ν = 0.598 ± 0.028 (95% confidence interval), that is indistinguishable from the 0.588 predicted for an excluded-volume random coil. The shaded region represents the 95% confidence intervals for future measurements, assuming that the errors about (log)R_G are normally distributed around the fitted relationship. Only the measurements for creatine kinase and angiotensin II fall outside this predictive interval, and, thus, only these measurements can be said to represent unambiguously significant deviations. Error bars indicate the reported experimental (i.e., standard) deviations of the sample. These were derived by using a variety of approaches and widely varying numbers of observations and therefore provide only an approximate indication of experimental precision.