Immobilized Proteins In Drug Discovery

National Institute On Aging

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Abstract

A new method for high throughput screening has been developed based upon the inclusion of the target receptors, transmembrane receptors (cellular, nuclear or mitochondrial) in a flowing chromatographic system. In this approach, the target protein is immobilized on a solid support and the support packed into a small column. The test chemicals are passed through the column, over the immobilized target, and the time that it takes for the compounds to pass from the beginning of the column to its end is directly related to the strength of interaction between the target and the compound, i.e. the binding affinity of the ligand-receptor complex. Using this method, complex chemical and biological mixtures can be rapidly sorted between compounds that interact and do not interact with the disease-related target. At the same time, the compounds that bind to the target are themselves rapidly sorted between low, medium and high affinity binders. Thus, the method quickly provides a large amount of data with high information content. We have developed columns containing various subtypes of the nicotinic receptor to screen tobacco smoke condensates. The general approach has also been expanded to the immobilization of SIRT6 protein onto the surface of an open tubular capillary and on the surface of magnetic beads. The SIRT6-MBs were used to extract small molecules that bind to SIRT6 from chemical and botanical mixtures. Additional SIRT6 activity assays were developed and optimized for the identification of SIRT6 activators and SIRT6 inhibitors. Using this approach, several novel SIRT6 activators have been identified from complex matrixes. And the pharmacological activity was used to create a pharmacophore model. More recently, rhBri2 was also immobilized onto the surface of magnetic beads. The Bri2-immobilized proteins was used to identify interacting proteins from a complex matrix.