Weak binders

It is important to emphasize that this lattice database is highly idealized compared to real databases. Unlike the lattice database, real databases cannot be treated as thermodynamic ensembles of protein-ligand complexes equilibrated at room temperature [33,34]. Two of the more straightforward reasons are mentioned here. First, real databases are inherently biased toward strong binders (K < 10 pM), because weak binders are difficult to crystallize and of lesser interest. Second, as mentioned above, real databases are not composed of a representative selection of proteins and ligands, and their compositions are biased toward peptide and peptidomimetic inhibitors and certain protein superfamilies. In contrast, because only one protein and four ligand types are used, the lattice database should have representative ligand compositions. 

In many respects the fragment approach is ideally suited to projects which have X-ray crystal structures available. The fragments are small and relatively weak binders, but they often only possess one pharmacophoric element that binds to a specific feature on the target. If this interaction is identified by X-ray structure determination, then project teams can propose specific plans which maintain that critical interaction, and ideally optimize binding through other vectors in their fragments. 

HCV IRES (Table 1, entry 6) A mass spectroscopy-based fragment approach was used to identify the weak binder 29 to the ribosome HA sub-domain of hepatitis C (HCV IRES). Conventional optimisation led to a sub-micromolar lead 30 [41]. 

In the combination strategy (Fig. 18.1), simple molecular fragments are screened, and those that bind to the target are then connected together to construct more potent compounds. Assuming that the dissociation constants of the compounds are multiplicative, linking even very weak binders can theoretically produce a potent inhibitor e.g. linking three... 

NMR screening methods can detect very weak binders, which shifts the measurable inhibition curve (Fig. 18.6) toward less complex molecules. So, it is theoretically more... 

Competition between components of a mixture becomes more likely as the hit rate and pool size increase. For a 10% hit rate, competing hits will be found in 26% of mixtures of ten compounds, but only 2.7% in mixtures of three [11]. Unless mixtures are deconvo-luted, competition can lead to false negatives when the competing compounds have different affinities. For screens using protein detection, competition can also produce false positives due to the additive effect of multiple weak binders. Competition can be controlled by limiting the size of mixtures and pooling dissimilar compounds to reduce the likelihood of two compounds binding at the same site. 

Case 3 No complexation. If the binding interaction is not strong enough there is no effect on the fluorescence. The fluorescence intensity of solutions of piperazinyl derivative 5a does not change on addition of metal ions. Ca(II) and Al(III), both very weak binders to the macrocyclic polyamines,(n) have no effect on the fluorescence of any anthrylazamacrocycle. Na+ also has no effect, and as shown in Figure 3.8 the titration profile of 5c with Zn(II) is virtually unaffected even by the presence of a 100,000-fold excess ofNa+ at pH 12. 

About 32 000 compounds were screened to identify compounds that bind non-covalently to RGS4 using the GPC spin column/ESI-MS methodology and 1720 compounds were identified to bind (including very weak binders) to RGS4 [15],... 

Fig. 2.16 GPC spin column binding assay of RGS4 and G alpha proteins with WY817 (MW 450 Da). Positive ion ESI mass spectra for compound WY817, a weak binder to RGS4 protein and non-binder to G alpha protein. A miniature P6 GPC spin column was used. (A) ESI-MS response for 250 pg of reference compound WY817 (no GPC spin column used), (B) ESI-MS response for GPC spin column (P6 gel, 1 cm long, 100 pL volume) eluate when 100 pg of WY817 were passed...

Zhao et al. implemented a structure-based docking protocol to narrow down 500 compounds from a database of 57 compounds in their pursuit of FKBPs inhibitors (89). A novel scaffold was designed using the information obtained from the binding mode analysis of a known weak binder. To avoid any scoring function shortcomings, three scoring functions were used to select the 500 compounds. Of these, 43 were synthesized and tested to identify one potent inhibitor in a mouse peripheral synthetic nerve model. 

The main challenge for any fragment screening method is the detection of weak binders. The methods commonly used to detect fragments can be broadly broken down into biophysical and functional assays. 

High-concentration screening (HCS) assays tend to be used less frequently than biophysical methods, but can be employed to good effect due to their high-throughput capacity. These assays require biochemical function of the protein, up front knowledge of biochemical activity, and it can be difficult to reliably detect weak binders in the millimolar range. Fluorescence correlation... 

The spectacular relationship between the nature of the X3 species and the promotion energy shows that the VBSCD is in fact a general model of the pseudo-Jahn-Teller effect (PJTE). A qualitative application of PJTE would predict all the X3 species to be transition-state structures that relax to the distorted X ---X-X and X-X— X entities. The VBSCD makes a distinction between strong binders, which form transition states, and weak binders that form stable intermediate clusters. Thus, the VBSCD model is in tune with the general observation that as one moves in the periodic table from strong binders to weak ones (e.g., metallic) matter changes from discrete molecules to extended delocalized clusters and/or lattices. The delocalized clusters of the strong binders are the transition states for chemical reactions. 

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