A physical model for PDZ-domain/peptide interactions

The PDZ domain is an interaction motif that recognizes and binds the C-terminal peptides of target proteins. PDZ domains are ubiquitous in nature and help assemble multiprotein complexes that control cellular organization and signaling cascades. We present an optimized energy function to predict the binding free energy (ΔΔG) of PDZ domain/peptide interactions computationally. Geometry-optimized models of PDZ domain/peptide interfaces were built using Rosetta, and protein and peptide side chain and backbone degrees of freedom are minimized simultaneously. Using leave-one-out cross-validation, Rosetta’s energy function is adjusted to reproduce experimentally determined ΔΔG values with a correlation coefficient of 0.66 and a standard deviation of 0.79 kcal mol−1. The energy function places an increased weight on hydrogen bonding interactions when compared to a previously developed method to analyze protein/protein interactions. Binding free enthalpies (ΔΔH) and entropies (ΔS) are predicted with reduced accuracies of R = 0.60 and R = 0.17, respectively. The computational method improves prediction of PDZ domain specificity from sequence and allows design of novel PDZ domain/peptide interactions

Investigations of cation-pi binding by cyclophane receptors in aqueous media

NOTE: Text or symbols not renderable in plain ASCII are indicated by [...]. Abstract is included in .pdf document. The binding properties of two new cyclophane receptors in aqueous media were explored. Replacement of two benzene rings of the host P with furans or thiophenes was expected to enhance cation-[pi] interactions, but significant improvements in the binding of positively-charged guests were not observed. Ab initio calculations provided a rationalization for the experimental findings and a better understanding of the cation-[pi] interaction. The binding in water of various guanidinium compounds to host P was also investigated. These molecules represented a new type of cationic guest for the receptor. Arginine was not measureably bound by P, but several alkylated guanidiniums were well bound, especially considering their high water solubility. Hexamethylguanidinium, a nonplanar molecule with [...] symmetry, was a particularly interesting guest which displayed enantioselective binding to P. In addition, some progess was achieved towards the synthesis of receptors that have either an amide or a carboxylate group attached to the rim of the cavity. The appended group is positioned such that it can form hydrogen bonds or electrostatic interactions with a bound guanidiniumn guest. Finally, exploratory work was carried out towards the synthesis of a cyclophane host that possesses a disulfide near the binding cavity. This receptor was designed to mimic the acetylcholine binding site of the nicotinic acetylcholine receptor. Although various disulfide containing macrocycles were synthesized, they were not sufficiently water soluble to permit studies of their binding properties

Molecular recognition in aqueous media. New binding studies provide further insights into the cation-π interaction and related phenomena

We describe a large number of binding studies in aqueous media designed to provide new insights into noncovalent binding interactions, especially the cation-π interaction. The studies include 7 different hosts, over 70 guests, and over 150 new binding constants. In addition to the now standard NMR methods, circular dichroism has proven to be an especially useful tool for determining aqueous binding constants. We have found that, in addition to the alkyliminium and tetraalkylammonium guests we have studied previously, sulfonium and guanidinium guests also show substantial cation-π effects. Bromination of the host greatly enhances its binding ability in a general fashion, primarily as a result of hydrophobic interactions. Addition of methoxy groups did not enhance binding, apparently as a result of a collapse of the host into a conformation that is not suitable for binding. Replacement of two benzene rings of the host by furans or thiophenes also did not enhance binding. Ab initio calculations provide a rationalization for this effect and suggest a clearer model for the cation-π interaction

Molecular recognition in aqueous media. New binding studies provide further insights into the cation-π interaction and related phenomena

We describe a large number of binding studies in aqueous media designed to provide new insights into noncovalent binding interactions, especially the cation-π interaction. The studies include 7 different hosts, over 70 guests, and over 150 new binding constants. In addition to the now standard NMR methods, circular dichroism has proven to be an especially useful tool for determining aqueous binding constants. We have found that, in addition to the alkyliminium and tetraalkylammonium guests we have studied previously, sulfonium and guanidinium guests also show substantial cation-π effects. Bromination of the host greatly enhances its binding ability in a general fashion, primarily as a result of hydrophobic interactions. Addition of methoxy groups did not enhance binding, apparently as a result of a collapse of the host into a conformation that is not suitable for binding. Replacement of two benzene rings of the host by furans or thiophenes also did not enhance binding. Ab initio calculations provide a rationalization for this effect and suggest a clearer model for the cation-π interaction

Intrinsically Disordered Regions Can Contribute Promiscuous Interactions to RNP Granule Assembly

Summary: Eukaryotic cells contain large RNA-protein assemblies referred to as RNP granules, whose assembly is promoted by both traditional protein interactions and intrinsically disordered protein domains. Using RNP granules as an example, we provide evidence for an assembly mechanism of large cellular structures wherein specific protein-protein or protein-RNA interactions act together with promiscuous interactions of intrinsically disordered regions (IDRs). This synergistic assembly mechanism illuminates RNP granule assembly and explains why many components of RNP granules, and other large dynamic assemblies, contain IDRs linked to specific protein-protein or protein-RNA interaction modules. We suggest assemblies based on combinations of specific interactions and promiscuous IDRs are common features of eukaryotic cells

A Derived Allosteric Switch Underlies the Evolution of Conditional Cooperativity between HOXA11 and FOXO1

Transcription factors (TFs) play multiple roles in development. Given this multifunctionality, it has been assumed that TFs are evolutionarily highly constrained. Here, we investigate the molecular mechanisms for the origin of a derived functional interaction between two TFs, HOXA11 and FOXO1. We have previously shown that the regulatory role of HOXA11 in mammalian endometrial stromal cells requires interaction with FOXO1, and that the physical interaction between these proteins evolved before their functional cooperativity. Here, we demonstrate that the derived functional cooperativity between HOXA11 and FOXO1 is due to derived allosteric regulation of HOXA11 by FOXO1. This study shows that TF function can evolve through changes affecting the functional output of a pre-existing protein complex

Exploring Symmetry as an Avenue to the Computational Design of Large Protein Domains

It has been demonstrated previously that symmetric, homodimeric proteins are energetically favored, which explains their abundance in nature. It has been proposed that such symmetric homodimers underwent gene duplication and fusion to evolve into protein topologies that have a symmetric arrangement of secondary structure elements“symmetric superfolds”. Here, the ROSETTA protein design software was used to computationally engineer a perfectly symmetric variant of imidazole glycerol phosphate synthase and its corresponding symmetric homodimer. The new protein, termed FLR, adopts the symmetric (βα)₈ TIM-barrel superfold. The protein is soluble and monomeric and exhibits two-fold symmetry not only in the arrangement of secondary structure elements but also in sequence and at atomic detail, as verified by crystallography. When cut in half, FLR dimerizes readily to form the symmetric homodimer. The successful computational design of FLR demonstrates progress in our understanding of the underlying principles of protein stability and presents an attractive strategy for the <i>in silico</i> construction of larger protein domains from smaller pieces