The biological efficacy of a small molecule drug candidate is coupled to its binding characteristics for its therapeutically relevant biomolecular target. The compound's most important binding characteristics include its affinity, binding site, and dissociation rate. Therefore, in any drug discovery program, considerable medicinal chemistry effort is expended to optimize these binding features of a drug candidate. In the early stages of the drug development process, progress towards improving a compound's binding features is typically followed using in vitro biochemical assays that measure a compound's effect on the conversion of one biological molecule into another, and other, orthogonal techniques that directly measure the binding characteristics of a lead compound for its receptor. These two methods are complementary, since direct protein-ligand binding assays provide the medicinal chemist with independent confirmation that activity observed in a biochemical assay correlates with specific binding to the target of interest, and that biochemical activity is not due to off-target binding, unwanted interaction with substrates or cofactors, or due to undesirable physical properties such as insolubility and target co-precipitation.

Techniques to directly characterize protein-ligand interactions play an increasingly vital role in the pharmaceutical discovery and development process. Direct binding assays are valuable not only to complement known techniques for determining the activity of ligands for well established classes of protein targets, but they are also critical for the pursuit of emerging drug targets that have no functional assay with which to evaluate potential ligands. In some cases, these techniques may be the only recourse for quantifying binding potency in a drug discovery program. For instance, advances in genome and proteome analysis are rapidly increasing the number of human and pathogen proteins identified as targets for small molecule therapy of human disease [1]. While these proteins may be synthesized and purified as targets for small-molecule therapy, many lack biochemical assays to discover and evaluate the binding properties of potential drug candidates, are only available in minute quantities, or lack endogenous ligands for affinity determination using competitive binding assays. Even classic targets with well established biochemical assays are yielding new avenues for therapy through non-traditional points of intersection along their reaction pathway, requiring sophisticated in vitro assays for the discovery and evaluation of new drug candidates. As an example, potential small-molecule therapeutics that bind to and prevent phosphorlyation of their basal kinase targets require complex, ''coupled'' assays to characterize their activities using purely biochemical means [2]. In instances where no such coupled assay exists, direct methods to measure protein-ligand binding characteristics are essential.

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