Neuronal and Cellular Signalling Lecture 9 Notes: The Nucleus, Signalling Cascades and Kinases Part 2 Tyrosine Kinases - Protein tyrosine kinases (PTKs) catalyse the transfer of phosphate groups from (usually) ATP to tyrosine residues in target (substrate) proteins - Phospho-tyrosine is a rare protein modification in mammalian cells (relatively to Ser/Thr phosphorylation). The extent of protein phosphorylation on Tyr (or Ser/Thr) is closely regulated by the opposing activities of kinases and protein phosphates. - Protein tyrosine kinases are a superfamily of 5 classes (sub-families): (a) AGC group (e.g. PKA, PKG, PKC, Rac, G-protein kinases) (b) CaMK group (kinases regulated by Ca2+/CaM) (c) CMGC group (Cyclin-dependent kinases, MAP, GSK3, Casein kinase) (d) PTK group (conventional protein tyrosine kinases, e.g. Src, Abl, Fak, PDGF and IR) (e) OPK (other protein kinases) - Tyrosine kinases can be cytosolic or integral membrane receptors. The general structure of cytosolic tyrosine kinases consists of a phosphorylated N-terminal (aas can be modified), SH3 and SH2 group, and C-terminal conserved kinase region. Integral membrane receptor tyrosine kinases consist of an N-terminal signal sequence (to embed the protein in the outer cell membrane), cysteine-rich region (usually involved in ligand binding) the membrane- spanning region (stretch of hydrophobic alpha helices), tyrosine kinase region and C-terminal tyrosine residues that are phosphorylated. - The primary aa sequence of PTKs have shared sequence motifs, allowing identification of a large family of human proteins, but relatively few are functionally understood. There are around 90 tyrosine kinases in the human genome. 58 are thought to be localised in the cell membrane (have TM structure) and have a receptor function coupled to Tyr kinase. 32 are cytoplasmic, have no TM a-helix but may be membrane-associated. - RTKs (receptor tyrosine kinases) function a variety of intracellular locations, including the intracellular face of the plasma membrane (C-terminal), at specialised regions of the cytoskeleton and in the nucleus. Their signalling functions at the plasma membrane are best understood. Examples of these growth factors which bind extracellular signals include: (a) EGF receptor (NEU/HER2/HER3) (b) Insulin receptor (c) PDGF (d) FGF (e) VEGF (f) Eph (g) M-CSF (macrophage colony stimulating factor) (h) NGF (nerve growth factor) - Receptors share a common structure. There is a lot of variability in the ligand-binding domain (IgG-like repeats necessary for ligand binding), single TM regions, sequence- conserved Tyr kinase region and C-terminal tail (functions to be phosphorylated). All RTKs have an extracellular binding domain with a ligand-binding site, a TM a-helix and cytoplasmic sequence with Tyr kinase activity. When the ligand (e.g. EGF) binds, this usually causes
dimerization of the receptor (activated state), activating the kinase domain and allowing phosphorylation of the receptor's tail. Ligand-RTK binding stimulates a signal transduction system which leads to control of cell physiology and gene expression (variety of consequences). -Ligand- This is involved in cell proliferation, differentiation, survival Inactive monomers and modulation of metabolism (i.e. involved in cancer). Other RTKs have been found in mutations that block the differentiation in particular cell types of C. elegans, mice and Drosophila. Active