Prediction and confirmation of a switch-like region within the N-terminal domain of hSIRT1.

Many proteins display conformational changes resulting from allosteric regulation. Often only a few residues are crucial in conveying these structural and functional allosteric changes. These regions that undergo a significant change in structure upon receiving an input signal, such as molecular recognition, are defined as switch-like regions. Identifying these key residues within switch-like regions can help elucidate the mechanism of allosteric regulation and provide guidance for synthetic regulation. In this study, we combine a novel computational workflow with biochemical methods to identify a switch-like region in the N-terminal domain of human SIRT1 (hSIRT1), a lysine deacetylase that plays important roles in regulating cellular pathways. Based on primary sequence, computational methods predicted a region between residues 186-193 in hSIRT1 to exhibit switch-like behavior. Mutations were then introduced in this region and the resulting mutants were tested for allosteric reactions to resveratrol, a known hSIRT1 allosteric regulator. After fine-tuning the mutations based on comparison of known secondary structures, we were able to pinpoint M193 as the residue essential for allosteric regulation, likely by communicating the allosteric signal. Mutation of this residue maintained enzyme activity but abolished allosteric regulation by resveratrol. Our findings suggest a method to predict switch-like regions in allosterically regulated enzymes based on the primary sequence. If further validated, this could be an efficient way to identify key residues in enzymes for therapeutic drug targeting and other applications.

Statistical potentials for RNA-protein interactions optimized by CMA-ES.

Characterizing RNA-protein interactions remains an important endeavor, complicated by the difficulty in obtaining the relevant structures. Evaluating model structures via statistical potentials is in principle straight-forward and effective. However, given the relatively small size of the existing learning set of RNA-protein complexes optimization of such potentials continues to be problematic. Notably, interaction-based statistical potentials have problems in addressing large RNA-protein complexes. In this study, we adopted a novel strategy with covariance matrix adaptation (CMA-ES) to calculate statistical potentials, successfully identifying native docking poses.

Logistic regression models to predict solvent accessible residues using sequence- and homology-based qualitative and quantitative descriptors applied to a domain-complete X-ray structure learning set.

A working example of relative solvent accessibility (RSA) prediction for proteins is presented. Novel logistic regression models with various qualitative descriptors that include amino acid type and quantitative descriptors that include 20- and six-term sequence entropy have been built and validated. A domain-complete learning set of over 1300 proteins is used to fit initial models with various sequence homology descriptors as well as query residue qualitative descriptors. Homology descriptors are derived from BLASTp sequence alignments, whereas the RSA values are determined directly from the crystal structure. The logistic regression models are fitted using dichotomous responses indicating buried or accessible solvent, with binary classifications obtained from the RSA values. The fitted models determine binary predictions of residue solvent accessibility with accuracies comparable to other less computationally intensive methods using the standard RSA threshold criteria 20 and 25% as solvent accessible. When an additional non-homology descriptor describing Lobanov-Galzitskaya residue disorder propensity is included, incremental improvements in accuracy are achieved with 25% threshold accuracies of 76.12 and 74.79% for the Manesh-215 and CASP(8+9) test sets, respectively. Moreover, the described software and the accompanying learning and validation sets allow students and researchers to explore the utility of RSA prediction with simple, physically intuitive models in any number of related applications.

Flexibility of BIV TAR-Tat: models of peptide binding.

A new approach in determining local residue flexibility from base-amino acid contact frequencies is applied to the twelve million lattice chains modeling BIV Tat peptide binding to TAR RNA fragment. Many of the resulting key features in flexibility correspond to RMSD calculations derived from a set of five NMR derived structures (X. Ye, R. A. Kumar, and D. J. Patel, Protein Data Bank: Database of three-dimensional structures determined from NMR (1996)) and binding studies of mutants (L. Chen and A. D. Frankel, Proc. Natl. Acad. Sci. USA 92, 5077-5081 (1995)). The lattice and RMSD calculations facilitate the identification of peptide hinge regions that can best utilize the introduction of Gly or other flexible residues. This approach for identifying potential sites amenable to substitution of more flexible residues to enhance peptide binding to RNA targets could be a useful design tool.