A non-receptor tyrosine kinase (nRTK) is a cytosolic enzyme that is responsible for catalysing the transfer of a phosphate group from a nucleoside triphosphate donor, such as ATP, to tyrosine residues in proteins. Non-receptor tyrosine kinases are a subgroup of protein family tyrosine kinases, enzymes that can transfer the phosphate group from ATP to a tyrosine residue of a protein (phosphorylation). These enzymes regulate many cellular functions by switching on or switching off other enzymes in a cell. Unlike the receptor tyrosine kinases (RTKs), the second subgroup of tyrosine kinases, the non-receptor tyrosine kinases are cytosolic enzymes. Thirty-two non-receptor tyrosine kinases have been identified in human cells (EC 2.7.10.2). Non-receptor tyrosine kinases regulate cell growth, proliferation, differentiation, adhesion, migration and apoptosis, and they are critical components in the regulation of the immune system.
Function The main function of nRTKs is their involvement in signal transduction in activated T- and B-cells in the immune system. Signaling by many receptors is dependent on nRTKs including T-cell receptors (TCR), B-cell receptors (BCR), IL-2 receptors (IL-2R), Ig receptors, erythropoietin (EpoR) and prolactin receptors. CD4 and CD8 receptors on T lymphocytes require for their signaling the Src family member Lck. When antigen binds to T-cell receptor, Lck becomes autophosphorylated and phosphorylates the zeta chain of the T-cell receptor, subsequently another nRTK, Zap70, binds to this T-cell receptor and then participates in downstream signaling events that mediate transcriptional activation of cytokine genes. Another Src family member Lyn is involved in signaling mediated by B-cell receptor. Lyn is activated by stimulation of B-cell receptor, which leads to the recruitment and phosphorylation of Zap70-related nRTK, Syk. Another nRTK, Btk, is also involved in signaling mediated by the B-cell receptor. Mutations in the Btk gene are responsible for X-linked agammaglobulinemia, a disease characterized by the lack of mature B-cells.
Structure Unlike receptor tyrosine kinases, nRTKs lack receptor-like features such as an extracellular ligand-binding domain and a transmembrane-spanning region. Most of the nRTKs are localized in the cytoplasm, but some nRTKs are anchored to the cell membrane through amino-terminal modification. These enzymes commonly have a modular construction and individual domains are joined together by flexible linker sequences. One important domain of nRTKs is the tyrosine kinase catalytic domain, which is about 275 residues in length. The structure of the catalytic domain can be divided into a small and a large lobe, where ATP binds to the small lobe and the protein substrate binds to the large lobe. Upon the binding of ATP and substrate to nRTKs, catalysis of phosphate transfer occurs in a cleft between these two lobes. It was found that nRTKs have some sequence preference around the target Tyr. For example, the Src preferred sequence is Glu–Glu/Asp–Ile–Tyr–Gly/Glu–Glu–Phe and Abl preferred sequence is Ile/Val–Tyr–Gly–Val–Leu/Val. Different preferred sequences around Tyr in Src and Abl suggest that these two types of nRTKs phosphorylates different targets. Non-receptor tyrosine kinases do not contain only a tyrosine kinase domain, nRTKs also possess domains that mediate protein-protein, protein-lipid, and protein-DNA interactions. One of the protein-protein interaction domains in nRTKs are the Src homology 2 (SH2) and 3 (SH3) domains. The longer SH2 domain (~100 residues) binds phosphotyrosine (P-Tyr) residues in a sequence-specific manner. The P-Tyr interacts with SH domain in a deep cleft, which cannot bind unphosphorylated Tyr. The SH3 domain is smaller (~60 residues) and binds proline-containing sequences capable of forming a polyproline type II helix. Some nRTKs without SH2 and SH3 domains possess some subfamily-specific domains used for protein-protein interactions. For example, specific domains that target enzymes to the cytoplasmic part of cytokine receptors (Jak family) or two domains: an integrin-binding domain and a focal adhesion-binding domain (Fak family). The nRTK Abl possess the SH2 and SH3 domains, but also possesses other domains for interactions: F actin–binding domain and a DNA-binding domain contains a nuclear localization signal and is found in both the nucleus and the cytoplasm. In addition to SH2 and SH3 domains, Btk/Tec subfamily of nRTKs possess another modular domain, a pleckstrin homology (PH) domain. These PH domains bind to phosphatidylinositol lipids that have been phosphorylated at particular positions on the head group. These enzymes can bind to activated signaling complexes at the membrane through PH domain interactions with phosphorylated phosphatidylinositol lipids.
Regulation The most common theme in nRTKs and RTK regulation is tyrosine phosphorylation. With few exceptions, phosphorylation of tyrosines in the activation loop of nRTKs leads to an increase in enzymatic activity. Activation loop phosphorylation occurs via trans-autophosphorylation or phosphorylation by different nRTKs. It is possible to negatively regulate kinase activity by the phosphorylation of tyrosines outside of the activation loop. Protein tyrosine phosphatases (PTPs) restore nRTKs to their basal state of activity. In some cases PTPs positively regulate nRTKs activity.
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