Comparing models of evolution for ordered and disordered proteins

Abstract

Most models of protein evolution are based upon proteins that form relatively rigid 3D structures. A significant fraction of proteins, the so-called disordered proteins, do not form rigid 3D structures and sample a broad conformational ensemble. Disordered proteins do not typically maintain long-range interactions, so the constraints on their evolution should be different than ordered proteins. To test this hypothesis, we developed and compared models of evolution for disordered and ordered proteins. Substitution matrices were constructed using the sequences of putative homologs for sets of experimentally characterized disordered and ordered proteins. Separate matrices, at three levels of sequence similarity (>85%, 85-60%, and 60-40%), were inferred for each type of protein structure. The substitution matrices for disordered and ordered proteins differed significantly at each level of sequence similarity. The disordered matrices reflected a greater likelihood of evolutionary changes, relative to the ordered matrices, and these changes involved nonconservative substitutions. Glutamic acid and asparagine were interesting exceptions to this result. Important differences between the substitutions that are accepted in disordered proteins relative to ordered proteins were also identified. In general, disordered proteins have fewer evolutionary constraints than ordered proteins. However, some residues like tryptophan and tyrosine are highly conserved in disordered proteins. This is due to their important role in forming protein-protein interfaces. Finally, the amino acid frequencies for disordered proteins, computed during the development of the matrices, were compared with amino acid frequencies for different categories of secondary structure in ordered proteins. The highest correlations were observed between the amino acid frequencies in disordered proteins and the solvent-exposed loops and turns of ordered proteins, supporting an emerging structural model for disordered proteins.

FIG. 1.

Iterative procedure used for constructing substitution matrices.

FIG. 2.

Amino acid frequencies (p_i) of residues in each matrix. The axis is sorted by average frequency of the residues in the D matrices. Shades of red are D matrices; shades of blue are O matrices. Bars indicate ±0.2%, which is within rounding error and slightly greater than the maximum standard deviation. D, disorder; O, order.

FIG. 3.

(A) Substitution matrices for ordered (O40, upper) and disordered (D40, lower) proteins at 40–60% sequence identity. Color shading indicates probability of substitutions being greater than expected by chance (blue) or less than expected (red). (B) Matrix showing the difference between D40 and O40. Color shading indicates greater frequency of substitutions in disorder (blue) or greater frequency in order (red). In all matrices, residues are ordered from most order promoting (green) to most disorder promoting (white) as shown in (Radivojac et al. 2007). D, disorder; O, order.

FIG. 4.

Residues that are most conserved are different between disordered and ordered proteins, and conservation is not determined by frequency. (A) Differences between the probability that an amino acid is conserved at a site (p_ii) for disorder and order at 40–60% identity for each of the 20 amino acids (i). Error bars indicate one standard deviation. (B) Scatter plot of the frequencies of each amino acid versus p_ii for disorder (filled diamonds) and order (open squares) at 40–60% identity.

FIG. 5.

Frequencies of amino acids in disordered proteins are most similar to the frequencies of amino acids in the exposed coils and turns of ordered proteins. The line indicates a one-to-one correspondence between frequency in order and disorder. (Information for disordered proteins is based upon the >85% similarity matrix and for ordered proteins is from Goldman et al. 1998.)

FIG. 6.

Scatter plot of the residue-normalized diagonal values, p_ii, for the eight secondary structure matrices and D85. The plot is ordered based upon increasing p_ii of the D85 matrix. Lines between data points from the same matrix are added for clarity. (Information for disordered proteins is based upon the >85% similarity matrix and for ordered proteins is from Goldman et al. 1998.). D, disorder; O, order.

FIG. 7.

Protein sequence alignment of the disordered p53 transactivation domain for seven closely related family members. The orange bar shows the position of the amphipathic helix that forms when p53 binds to the ubiquitin ligase, MDM2. The green bar shows the position of the amphipathic helix that forms when p53 binds to RPA70.