doi:10.1016/j.chemphys.2004.05.008 
Copyright © 2004 Elsevier B.V. All rights reserved.
Guiding the search for a protein's maximum rate of folding
Yongjin Zhua, 1, Xiaoran Fua, b, 1, Ting Wanga, 1, Atsuo Tamurac, Shoji Takadac, Jeffery G. Saven
, 
, a, b and Feng Gai, , a
a Department of Chemistry, University of Pennsylvania, 231 S. 34th St., Philadelphia, PA 19104, USA
b Makineni Theoretical Laboratories, University of Pennsylvania, Philadelphia, PA 19104, USA
c Graduate School of Science and Technology, Kobe University, Rokkodai, Nada, Kobe 657-8501, Japan
Received 1 February 2004;
accepted 3 May 2004.
Available online 1 June 2004.
References and further reading may be available for this article. To view references and further reading you must
purchase this article.
Abstract
Many simple, single-domain proteins fold via first order kinetics, indicative of a single, dominant free energy barrier. Because folding usually involves the burial of hydrophobic side chains, the acquisition of native structure may be associated with a decrease in the heat capacity of the system. If the transition state ensemble involves the burial of hydrophobic residues, the folding rates show a well-known concave downward dependence on temperature, exhibiting a maximum folding rate with respect to temperature. Within the framework of transition state theory, the maximum folding rate for a specific native structure depends simply on the entropic barrier as well as the heat capacity of activation. The latter is related to the mean hydrophobicity when the protein is largely unfrustrated with regard to its stabilizing interactions. As an example, here we show that the maximum folding rate of the three-helix bundle structure of 1prb7–53, the GA module of an albumin binding domain, can indeed be fine-tuned using computational design methods to identify and design structurally consistent mutations that modulate its hydrophobic content. Specifically, we find that the logarithm of the maximal folding rate depends linearly on the mean hydrophobic content of the designed sequences, where faster folding correlates with higher mean hydrophobicity.
Fig. 1. The NMR solution structure of 1prb (the 6 N-terminal residues are not shown) [25].
Fig. 2. Calculated amino acid probability distributions at selected positions. The mutation positions in numbers and the wild type residues in one-letter codes are indicated at upper-right corner of each panel. The double mutation is chosen based on the same results.
Fig. 3. Temperature dependent mean residue ellipticities at 222 nm of 1prb7–53 and its mutants in 0 M (
), 3 M (□), 5 M (
), and 7 M (×) GuHCl D2O solution (50 mM phosphate buffer, pH* 7.0). Solid lines are fits according to the method described in the text. The resulting thermodynamic parameters are summarized in Table 2.
Fig. 4. (a) IR spectra of 1prb7–53 in D2O (50 mM phosphate buffer, pH* 7.0) corresponding to different temperatures, as indicated (the data have been offset for clarity). These spectra were modeled (solid lines) globally by five Gaussians. The bands that make up the fit for the 3.9 °C spectrum are shown (ν1=1609.8 cm−1, Δν1=24.2 cm−1; ν2=1632.6 cm−1, Δν2=25.1 cm−1; ν3=1643.7 cm−1, Δν3=31.9 cm−1; ν4=1654.1 cm−1, Δν4=19.6 cm−1; ν5=1667.3 cm−1, Δν5=29.5 cm−1; where ν=band position and Δν=full width at half maximum). The band at 1610 cm−1 arises from amino acid sidechains, whereas the remaining four bands are due to amide C=Os. (b) Relative band areas of these bands as a function of temperature.
Fig. 5. A representative relaxation kinetic trace of 1prb7–53 corresponding to a T-jump of 83–96.2 °C. The probing frequency is 1630 cm−1. The solid line is the best fit to the following equation: ΔOD(t)=A*[1−B*exp(−t/τ)] with A=−6.6, B=0.55 and τ=3.4 μs.
Fig. 6. Arrenhius plot of the observed relaxation rate constant () as well as the folding () and unfolding (□) rate constants for 1prb7–53 and its mutants, as indicated. Lines are fits to Eqs. ((1) and (2)). Reciprocal temperature corresponds to the final temperature of the laser initiated T-jump.
Fig. 7. Logarithm of the maximum folding rates of 1prb7–53 and its mutants vs. (a) the mean hydrophobicity and (b) the calculated environmental energy (ΔEenv,ΔEenv=Eenv(mutant)−Eenv(w.t.)).
Table 2. Unfolding thermodynamic parameters in the absence of denaturant determined from CD spectroscopy for the wild type and five mutants of 1prb7–53 in D2O (50 mM phosphate buffer, pH*=7), also listed are the minimum folding times of these proteins and the corresponding Tmax
Abstract
Purchase PDF (755 K)
E-mail Article
Add to my Quick Links
Cited By in Scopus (26)
Purchase PDF (210 K)
env=