Protein kinase

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A protein kinase is an enzyme that can transfer a phosphate group from a donor molecule (usually ATP) to an amino acid residue of a protein. This mechanism is used in signal transduction for the regulation of enzymes; such a phosphorylation can activate (or inhibit) the enzymatic activity of an enzyme. While most protein kinases are specialized on a single kind of amino acid residue, some of them have a dual kinase activity (they can phosphorylate two different kinds of amino acids). Protein kinases can be regulated by

  • Activator proteins
  • Inhibitor proteins
    • Pseudosubstrates
    • Autoinhibition (a part of the protein kinase mimics a pseudosubstrate)
  • Ligands binding to regulatory subunits
  • Cofactors / second messenger
  • Phosphorylation in the active center (intrasterical regulation) by
    • other protein kinases (trans-phosphorylation)
    • itself (cis-phosphorylation/autophosphorylation)
  • Localization in the cell

Serine/threonine-specific protein kinases

Serine/threonine protein kinases phosphorylate the OH group of serine or threonine (which have quite similar residues). These protein kinases can be regulated by:

These kinases do not have a similar consensus sequence; there is no common "target sequence" to be phosphorylated. As the substrate to be phosphorylated will align to the kinase by several key amino acids (usually by hydrophopic forces and ion bonds), a kinase is usually spefic not to a single substrate, but to a whole "substrate family" with common properties. The kinases are usually inactivated by a pseudosubstrate that binds to the kinase similar to a real substrate, but lacks the amino acid to phosphorylated. Its removal will activate the kinase.
The catalytic domain of the kinases is highly conserved.

Phosphorylase kinase

Phosphorylase kinase was the first Ser/Thr protein kinase to be discovered (in 1959 by Krebs et al.).

Protein kinase A

The protein kinase A consists of two domains, a small domain with several β sheet structures, and a larger domain containing several α helices. In the catalytic cleft between the domains (or lobes), the the substrate and ATP binding sites are located. On ATP and substrate binding, the two lobes rotate so the phosphate group of the ATP and the target amino acid of the substrate move into the right position for the catalytic reaction to take place.

Regulation

Protein kinase A has several function within the cell, including regulation of glycogen-, sugar- and lipid metabolism. It is controlled by cAMP; in the absence of cAMP, the kinase is a tetramer of each two regulatory and two catalytic subunits (R2C2), with the regulatory subints blocking the catalytic center of the catalytic subunits. Binding of cAMP to the regulatory subunit leads to the dissociation of active RC-dimers. Also, the catalytic subunit itself can be regulated by phosphorylation.
Downregulation of protein kinase A is done by a feedback mechanism. One of the substrates that is activated by the kinase is a phosphodiestrase, which converts cAMP to AMP, thus reducing the amount of cAMP that can activate protein kinase A.

Protein kinase C

Protein kinase C actually describes a family of protein kinases that need Ca2+, diacylglycerole, and a phospholipid like phosphatidylcholine for activation. This shows that protein kinase C is activated through the same signal transduction pathway as phospholipase C. There have been at least twelve members of the proteine kinase C family identified in mammals by their high sequence homology. The protein kinase C usually means the protein kinase Cα enzyme.

Structure and regulation

Protein kinase C enzymes consist of a N-terminal regulatory domain, and a C-terminal catalytic domain. The kinases are inactive in the absence of activating agents by autoinhibition of the regulatory domain. They can be activated tumor promotors like tetradecanoyle-phorbole-acetate (or TPA), or by the cofactors Ca2+, diacylglycerole and a phospholipid. The common linear structure of protein kinase C enzymes is :

N - pseudosubstrate - TPA-binding - (Ca2+-binding) - ATP-binding - substrate-binding - C 

Upon activation, protein kinase C enzymes are translocated to the plasma membrane by RACK proteins (membrane-bound receptor for activated protein kinase C proteins). The protein kinase C enzymes are known for their long-term activation; they are still activated after the original activation signal or the Ca2+-wave are already gone. This is presumably achieved by the protuction of diacylglycerole from phosphatidylcholine by a phospholipase; fatty acids may also play a role in long-term activation.

Function

The consensus sequence of protein kinase C enzymes is similar to that of protein kinase A, as it contains basic amino acids close to the Ser/Thr to be phosphorylated. Their substrates are MARCKS proteins, MAP kinase, transcription factor inhibitor IXB, the vitamin D3 receptor VDR, Raf kinase, calpaine, and the EGF receptor.

Ca2+/calmodulin-dependent protein kinases

Also called CaM kinases, these kinases are primarily regulated by the Ca2+/calmodulin complex. A remakably property of these kinases is a memory effect on activation. There are two types of CaM kinases:

  1. Specialized CaM kinases. An example is the Myosine Light Chain Kinase (MLCK) that phosphorylates myosine, causing muscles to contract.
  2. Multifunctional CaM kinases. Also collectively called CaM kinase II, they are engaged in a multitude of processes, like neurotransmitter secretion, transcription factor regulation, and glycogen metabolism.

Structure and autoregulation

The Cam kinases consist of a N-terminal catalytic domain, a regulatory domain, and an associative domain. In the absence of Ca2+/calmodulin, the catalytic domain is autoinhibited by the regulatory domain, which conteins a pseudosubstrate sequence. Several Cam kinases aggregate to a homooligomer or heterooligomer. On activation by Ca2+/calmodulin, the activated CaM kinases autophosphorylate each other in an intermolecular reaction. This has two effects:

  1. It increases the affinity for the calmodulin complex, prolonging the time the kinase is active.
  2. The phosphorylated kinase complex remains active for a while even after the calmodulin complex has dissociated from the kinase complex, prolonging the active state even further.

MAP kinases

Mos/Raf kinases

Tyrosine-specific protein kinases

Tyrosine-specific protein kinases are, like serine/threonine-specific kinases, used in signal transguction. They act primarily as growth factor receptors, for example, the platlet derived growth factor, epidermal growth factor, transforming growth factor, insulin and insulin-like growth factor, interleukines, and tumor necrosis factor.

Receptor tyrosine kinases

These kinases consist of a transmembrane receptor with a tyrosine kinase domain integrated in the cytoplasmic domain. They play an important role in the regulation of cell division, cell differentiation, and morphogenesis. There are more than 50 known receptor tyrosine kinases in mammals.

Structure

The extracellular domain serves as the ligand receptor. It can be a sepatate unit that is attached to the rest of the receptor via a disulfide bond. The same mechanism can be used to bind two receptors together to form a homo- or heterodimer. The transmembrane element is a single α helix. The intracellular or cytoplasmic domain contains the (highly conserved) kinase activity, as well as several regulatory functions.

Regulation

The ligand binding causes two reactions:

  1. The dimerisation of two monomeric receptor kinases, or the stabelization of a loose dimer. Many ligands of receptor tyrosine kinases are multivalent. Some tyrosine receptor kinases (e.g., the platelet derived growth factor receptor) can build heterodimers with other similar, but not equal kinases of that subfamily, allowing a much varied response to the extracellular signal.
  2. The 'trans-autophosphorylation (phosphorylation by the other kinase in the dimer) of the kinase.

The autophosphorylation causes the two subdomains of the intrinsic kinase to shift, opening the kinase domain for ATP binding. In the inactive form, the kinase subdomains are aligned in a way that ATP cannot reach the catalytic center of the kinase. When several amino acids suited for phosphorylation are present in the kinase domain (e.g., the insulin-like growth factor receptor), the activity of the kinase can increase with the number of phosphorylated amino acids; in this case, the first phosphorylation is said to be a cis-autophosphorylation, switching the kinase form "off" to "standby".

Signal transduction

The active tyrosine kinase phosphorylates specific target proteins, which are often enzymes themselves. An important target is the ras protein signal transduction chain.

Histidine-specific protein kinases

Aspartic acid/glutamic acid-specific protein kinases