Intrinsically Disordered Proteins Display No Preference for Chaperone Binding In Vivo

Intrinsically disordered/unstructured proteins (IDPs) are extremely sensitive to proteolysis in vitro, but show no enhanced degradation rates in vivo. Their existence and functioning may be explained if IDPs are preferentially associated with chaperones in the cell, which may offer protection against degradation by proteases. To test this inference, we took pairwise interaction data from high-throughput interaction studies and analyzed to see if predicted disorder correlates with the tendency of chaperone binding by proteins. Our major finding is that disorder predicted by the IUPred algorithm actually shows negative correlation with chaperone binding in E. coli, S. cerevisiae, and metazoa species. Since predicted disorder positively correlates with the tendency of partner binding in the interactome, the difference between the disorder of chaperone-binding and non-binding proteins is even more pronounced if normalized to their overall tendency to be involved in pairwise protein–protein interactions. We argue that chaperone binding is primarily required for folding of globular proteins, as reflected in an increased preference for chaperones of proteins in which at least one Pfam domain exists. In terms of the functional consequences of chaperone binding of mostly disordered proteins, we suggest that its primary reason is not the assistance of folding, but promotion of assembly with partners. In support of this conclusion, we show that IDPs that bind chaperones also tend to bind other proteins

On computational approaches for size-and-shape distributions from sedimentation velocity analytical ultracentrifugation

Sedimentation velocity analytical ultracentrifugation has become a very popular technique to study size distributions and interactions of macromolecules. Recently, a method termed two-dimensional spectrum analysis (2DSA) for the determination of size-and-shape distributions was described by Demeler and colleagues (Eur Biophys J 2009). It is based on novel ideas conceived for fitting the integral equations of the size-and-shape distribution to experimental data, illustrated with an example but provided without proof of the principle of the algorithm. In the present work, we examine the 2DSA algorithm by comparison with the mathematical reference frame and simple well-known numerical concepts for solving Fredholm integral equations, and test the key assumptions underlying the 2DSA method in an example application. While the 2DSA appears computationally excessively wasteful, key elements also appear to be in conflict with mathematical results. This raises doubts about the correctness of the results from 2DSA analysis

Residual structures, conformational fluctuations, and electrostatic interactions in the synergistic folding of two intrinsically disordered proteins

To understand the interplay of residual structures and conformational fluctuations in the interaction of intrinsically disordered proteins (IDPs), we first combined implicit solvent and replica exchange sampling to calculate atomistic disordered ensembles of the nuclear co-activator binding domain (NCBD) of transcription coactivator CBP and the activation domain of the p160 steroid receptor coactivator ACTR. The calculated ensembles are in quantitative agreement with NMRderived residue helicity and recapitulate the experimental observation that, while free ACTR largely lacks residual secondary structures, free NCBD is a molten globule with a helical content similar to that in the folded complex. Detailed conformational analysis reveals that free NCBD has an inherent ability to substantially sample all the helix configurations that have been previously observed either unbound or in complexes. Intriguingly, further high-temperature unbinding and unfolding simulations in implicit and explicit solvents emphasize the importance of conformational fluctuations in synergistic folding of NCBD with ACTR. A balance between preformed elements and conformational fluctuations appears necessary to allow NCBD to interact with different targets and fold into alternative conformations. Together with previous topology-based modeling and existing experimental data, the current simulations strongly support an ‘‘extended conformational selection’’ synergistic folding mechanism that involves a key intermediate state stabilized by interaction between the C-terminal helices of NCBD and ACTR. In addition, the atomistic simulations reveal the role of long-range as well as short-range electrostatic interactions in cooperating with readily fluctuating residual structures, which might enhance the encounter rate and promote efficient folding upon encounter for facile binding and folding interactions of IDPs. Thus, the current study not only provides a consistent mechanistic understanding of the NCBD/ACTR interaction, but also helps establish a multi-scale molecular modeling framework for understanding the structure, interaction, and regulation of IDPs in general