Crystal Structures of IRAK-4 Kinase in Complex with Inhibitors: A Serine/Threonine Kinase with Tyrosine as a Gatekeeper

Abstract

Interleukin-1 (IL-1) receptor-associated kinase-4 (IRAK-4) is a serine/threonine kinase that plays an essential role in signal transduction by Toll/IL-1 receptors (TIRs). Here, we report the crystal structures of the phosphorylated human IRAK-4 kinase domain in complex with a potent inhibitor and with staurosporine to 2.0 and 2.2 A, respectively. The structures reveal that IRAK-4 has a unique tyrosine gatekeeper residue that interacts with the conserved glutamate from helix alphaC. Consequently, helix alphaC is "pulled in" to maintain the active orientation, and the usual pre-existing hydrophobic back pocket of the ATP-binding site is abolished. The peptide substrate-binding site is more open when compared with other protein kinases due to a marked movement of helix alphaG. The pattern of phosphate ligand interactions in the activation loop bears a close resemblance to that of a tyrosine kinase. Our results provide insights into IRAK-4 function and the design of selective inhibitors.

Interleukin 1/Toll-like Receptor-induced Autophosphorylation Activates Interleukin 1 Receptor-associated Kinase 4 and Controls Cytokine Induction in a Cell Type-specific Manner

“…Previously, Cheng et al (10) identified Thr-342, Thr-345, and Ser-346 by MS analysis, and other groups identified these residues by crystallography (19,22). In this report, we present several lines of evidence to support the hypothesis that autophosphorylation of IRAK4 is required for the full activation and subsequent signaling events in a cell type-specific manner.…”

“…We also examined primary human monocytes from healthy volunteers, because we were interested in innate immune cells where IRAK4 activity plays an important role as an inflammatory mediator. We used compound 1, a previously characterized inhibitor of IRAK4 kinase for our studies on the pharmacology of IRAK4 in human cells (19). This compound is highly potent against IRAK4 (IC 50 ϭ 2 nM at K m ϭ 300 M ATP) (22) and to a lesser extent IRAK1 (IC 50 ϭ 15 nM at K m ϭ 35 M ATP).…”

“…Importantly, our data using the IRAK4 inhibitor compound 1 also support that IRAK4 phosphorylation is an autophosphorylation event and is associated with cytokine induction in a cell type-specific manner. The inhibitor used has an IC 50 for IRAK4 of 2 nM and for IRAK1 of 15 nM (19), making it difficult to definitively assess the contribution of either kinase on cellular signaling. However, assessment of ATP-binding site occupancy by the inhibitor in the cell shows that at the concentrations tested (10 M), IRAK4 is 90% occupied, whereas IRAK1 is only 55% occupied by the inhibitor, indicating that most of the effects are derived from inhibition of IRAK4 (see supplemental Table S1).…”

Cutting Edge: IL-1 Receptor-Associated Kinase 4 Structures Reveal Novel Features and Multiple Conformations

“…After submission of this manuscript, Wang et al (24) published the crystal structure of IRAK4 kinase domain complexed with staurosporine and a benzimidazole inhibitor.…”

An Innate Immunity Signaling Process Suppresses Macrophage ABCA1 Expression through IRAK-1-Mediated Downregulation of Retinoic Acid Receptor α and NFATc2

“…As demonstrated previously, the kinase domain of IRAK-1 may adopt a tertiary structure that resembles the cyclin-dependent kinase. (28,58). Consequently, IRAK-1 preferentially phosphorylates substrate containing Ser/Thr-Pro-rich motifs.…”