Biosensor-Based Approach to the Identification of Protein Kinase Ligands with Dual-Site Modes of Action

TY - JOUR

T1 - Biosensor-Based Approach to the Identification of Protein Kinase Ligands with Dual-Site Modes of Action

AU - Navratilova, Iva

AU - Macdonald, Graeme

AU - Robinson, Colin

AU - Hughes, Samantha

AU - Mathias, John

AU - Phillips, Chris

AU - Cook, Andrew

PY - 2012/2

Y1 - 2012/2

N2 - The authors have used a surface plasmon resonance (SPR)-based biosensor approach to identify and characterize compounds with a unique binding mode to protein kinases. Biacore was used to characterize hits from an enzymatic high-throughput screen of the Tec family tyrosine kinase, IL2-inducible T cell kinase (ITK). Complex binding kinetics was observed for some compounds, which led to identification of compounds that bound simultaneously at both the adenosine triphosphate (ATP) binding site and a second, allosteric site on ITK. The presence of the second binding site was confirmed by X-ray crystallography. The second site is located in the N-terminal lobe of the protein kinase catalytic domain, adjacent to but distinct from the ATP site. To enable rapid optimization of binding properties, a competition-based Biacore assay has been developed to successfully identify second site noncompetitive binders that have been confirmed by X-ray crystallographic studies. The authors have found that SPR technology is a key method for rapid identification of compounds with dual-site modes of action.

AB - The authors have used a surface plasmon resonance (SPR)-based biosensor approach to identify and characterize compounds with a unique binding mode to protein kinases. Biacore was used to characterize hits from an enzymatic high-throughput screen of the Tec family tyrosine kinase, IL2-inducible T cell kinase (ITK). Complex binding kinetics was observed for some compounds, which led to identification of compounds that bound simultaneously at both the adenosine triphosphate (ATP) binding site and a second, allosteric site on ITK. The presence of the second binding site was confirmed by X-ray crystallography. The second site is located in the N-terminal lobe of the protein kinase catalytic domain, adjacent to but distinct from the ATP site. To enable rapid optimization of binding properties, a competition-based Biacore assay has been developed to successfully identify second site noncompetitive binders that have been confirmed by X-ray crystallographic studies. The authors have found that SPR technology is a key method for rapid identification of compounds with dual-site modes of action.

U2 - 10.1177/1087057111422746

DO - 10.1177/1087057111422746

M3 - Article

C2 - 22068706

SN - 1087-0571

VL - 17

SP - 183

EP - 193

JO - Journal of Biomolecular Screening

JF - Journal of Biomolecular Screening

IS - 2

ER -