Design and Nuclear Magnetic Resonance (NMR) Structure Determination of the Second Extracellular Immunoglobulin Tyrosine Kinase A (TrkAlg2) Domain Construct for Binding Site Elucidation in Drug Discovery

作者：Shoemark Debbie K; Williams Christopher; Fahey Mark S; Watson Judy J; Tyler Sue J; Scoltock Simon J; Ellis Rosamund Z; Wickenden Elaine; Burton Antony J; Hemmings Jennifer L; Bailey Christopher D; Dawbarn David; Jane David E; Willis Christine L; Sessions Richard B; Allen Shelley J; Crump Matthew P^*

来源：Journal of Medicinal Chemistry, 2015, 58(2): 767-777.

DOI：10.1021/jm501307e

The tyrosine kinase A (TrkA) receptor is a validated therapeutic intervention point for a wide range of conditions. TrkA activation by nerve growth factor (NGF) binding the second extracellular immunoglobulin (TrkAIg(2)) domain triggers intracellular signaling cascades. In the periphery, this promotes the pain phenotype and, in the brain, cell survival or differentiation. Reproducible structural information and detailed validation of proteinligand interactions aid drug discovery. However, the isolated TrkAIg(2) domain crystallizes as a beta-strand-swapped dimer in the absence of NGF, occluding the binding surface. Here we report the design and structural validation by nuclear magnetic resonance spectroscopy of the first stable, biologically active construct of the TrkAIg(2) domain for binding site confirmation. Our structure closely mimics the wild-type fold of TrkAIg(2) in complex with NGF (1WWW.pdb), and the H-1-N-15 correlation spectra confirm that both NGF and a competing small molecule interact at the known binding interface in solution.

• 出版日期2015-1-22