This chimera will serve as a tool for a deeper understanding of the relationship between structure and thermodynamics of binding using a combination of NMR, stability and site-directed mutagenesis studies, which can lead to an effective strategy for ligand design.
Here, we elucidate detailed events of substrate binding, through a combination of crystallography, molecular dynamics, site-directed mutagenesis, biochemical/biophysical assays, and the design and synthesis of novel ligands.
Modeling showed that the Cys4 binding pocket created by displaying cysteines on a β-barrel scaffold in these variants was too large to allow simultaneous coordination of Zn2+ by all four cysteines and suggested that a binding site consisting of a combination of cysteines and histidines might provide a better Zn2+ binding site.
Increased protection from hydrogen-deuterium exchange in these regions could possibly indicate involvement of these Spy residues in the interaction with Im7 or could imply the folding of these flexible regions upon client binding, or a combination of both.
To locate places where a co-binding of a TF combination may happen, we identified genomic regions that contain at least two binding sites of main factors of interest in the ChIP-seq data.
These results suggest that client binding may occur over large regions of Spy, that Spy might undergo significant conformational changes upon client binding, or a combination of both (Quan et al., 2011).
Peak-motifs facilitates the discovery of binding motifs using a combination of several algorithms at a time, and it detects not only the strongest motif but also secondary ones, providing valuable information concerning cofactors, and mechanism of function for TFs (88).
Using an Epstein-Barr virus-derived episomal vector, antibody libraries are displayed as whole IgG molecules on the cell surface and screened for specific antigen binding by a combination of magnetic beads and fluorescence-activated cell sorting.
Importin beta has a lesser effect than RanBP7 on sperm chromatin decondensation and H1 binding, while a combination of RanBP7/importin beta is no more effective than RanBP7 alone.
Given the effect on both parameters it is not clear whether the W378A mutant is compromised in catalytic activity, has a severe defect in substrate binding, or a combination of the two.
