Ligand-based NMR screens

Binding energies are also routinely used to rank different ligands from a compound library after being docked to a protein target. The virtual or in silico screening of a library composed of thousands of theoretical compounds can be accomplished in a day with minimal cost [47 9]. Thus, a virtual screen can significantly accelerate the hit identification and optimization process while 

NMR ligand affinity screen and protein molecular weight is not a limiting factor 

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Table I Advantages and disadvantages of target- and ligand-based NMR screening methods...

In fact, higher molecular-weight proteins enhance the observation of abinding event in a ligand-based NMR screen. All of these characteristics make ligand-based NMR screens a routinely used drug discovery technique. 

We have used a highly structure-driven approach composed of fragment-based NMR screening, X-ray crystallography, and structure-assisted chemistry to develop a first-in-class clinical candidate as a potential proof-of-concept for the inhibition of BACE-1 in AD. Crucial to this achievement was the initial identification of a ligand-efficient isothiourea fragment and its X-ray crystal structure, which revealed an extensive H-bond network with the two active site aspartates. This interaction was unprecedented in the aspartic acid protease field when we discovered it several... 

Dalvit C (2008) Theoretical analysis of the competition ligand-based NMR experiments and selected applications to fragment screening and binding constant measurements. Concepts Magn Reson A 32A 341-372... 

C. Dalvit, Theoretical Analysis of the Competition Ligand-Based NMR Experiments and Selected Applications to Fragment Screening and Binding Constant Measurements , Concepts Magn. Reson., A, 2008, 32A, 341. 

Fig. 1 Ligand-based NMR methods for fragment library screening, (a) 1D H NMR controi spectrum of reference compound, (b) STD NMR spectrum of reference compound in the presence of the target protein. Only resonances of atoms which contact the protein are present in the STD spectrum (c) c-STD of reference compound and known competitor in the presence of the target protein. The STD signal of reference compound decreases because the known competitor with higher affinity displaces it from the binding site...

In this chapter we present the detailed setup of ligand-based NMR experiments and an NMR-based functional assay using a 30 kDa enzyme protein that makes bacteria resistant to a broad range of beta-lactam antibiotics. STD, c-STD, and STDD were used for fragment screening and an NMR-based functional assay that monitors the hydrolysis of ampicillin was used for functional validation of fragment hits with weak affinities/activities. 

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