Structure of the PH domain and Btk motif from Bruton's tyrosine kinase: molecular explanations for X-linked agammaglobulinaemia

Abstract

Bruton's tyrosine kinase (Btk) is an enzyme which is involved in maturation of B cells. It is a target for mutations causing X‐linked agammaglobulinaemia (XLA) in man. We have determined the structure of the N‐terminal part of Btk by X‐ray crystallography at 1.6 Å resolution. This part of the kinase contains a pleckstrin homology (PH) domain and a Btk motif. The structure of the PH domain is similar to those published previously: a seven‐stranded bent β‐sheet with a C‐terminal α‐helix. Individual point mutations within the Btk PH domain which cause XLA can be classified as either structural or functional in the light of the three‐dimensional structure and biochemical data. All functional mutations cluster into the positively charged end of the molecule around the predicted binding site for phosphatidylinositol lipids. It is likely that these mutations inactivate the Btk pathway in cell signalling by reducing its affinity for inositol phosphates, which causes a failure in translocation of the kinase to the cell membrane. A small number of signalling proteins contain a Btk motif that always follows a PH domain in the sequence. This small module has a novel fold which is held together by a zinc ion bound by three conserved cysteines and a histidine. The Btk motif packs against the second half of the β‐sheet of the PH domain, forming a close contact with it. Our structure opens up new ways to study the role of the PH domain and Btk motif in the cellular function of Btk and the molecular basis of its dysfunction in XLA patients.

Akt activation by growth factors is a multiple-step process: the role of the PH domain

“…This structural model is supported by ®ndings showing that all the mutations in the predicted phospholipid binding pocket a ect PH domain function. Moreover, the Akt residues R15, R25 and E40, which are predicted to lie in the phospholipid binding pocket of Akt, structurally align with the corresponding residues K12, R28 and E41 in the recently published structure of the Btk PH domain (Hyvonen and Saraste, 1997).…”

“…Residues R15, R25 and E40 are pointing into the proposed pocket. Interestingly, the recently published crystal structure of the PH domain of Btk shows that residues K12, R28, and E41 (which correspond to residues R15, R25 and E40 in Akt) are also located in a well de®ned lipid binding pocket (Hyvonen and Saraste, 1997). Residues in the positive surface patch were selected for mutation.…”

Tec Kinase Signaling in T Cells Is Regulated by Phosphatidylinositol 3-Kinase and the Tec Pleckstrin Homology Domain

“…These studies have strongly implicated the PH domain in membrane targeting of Tec kinases, thus establishing the importance of Tec kinase recruitment to the site of an activated receptor, and indicating the role of PI3K in Tec kinase activation (32,36). The vast majority of these studies have been performed on Btk, owing in part to the fact that a three-dimensional structure of the Btk PH domain has been determined (45). These data have demonstrated that substitutions at residue R28 in Btk (comparable to R29 in Tec) dramatically reduce binding of the Btk PH domain to D3 phosphoinositides, whereas substitution of FIGURE 2.…”

“…Specifically, the glutamic acid residue at position 41 in Btk (42 in other Tec kinases) is thought to be too far away from the inositol-binding pocket to directly interact with the inositol phosphate moiety of phosphatidylinositol-3,4,5-trisphosphate (PtdInsP3). Instead, this residue may enhance Btk binding to the membrane by binding other polar head groups of phospholipids in the membrane or, alternatively, may stabilize a putative dimerization of the PH domain by creating a more favorable interaction with the glutamic acid residue at position 45 in the dimer interface (45,55). Interestingly, the other Tec family PH domains, including Tec and Bmx, have an arginine or lysine residue at position 44 or 45; thus, substitution of E42 with lysine may, in fact, destabilize the putative dimer structure.…”

BTKbase: the mutation database for X-linked agammaglobulinemia

“…The three-dimensional structure has been determined for the PH domain and the first half of the TH domain [Hyvönen and Saraste, 1997], the SH3 domain [Hansson et al, 1998], and the kinase domain [Mao et al, 2001]. The SH2 domain structure has been modeled [Vihinen et al, 1994a].…”