A-myristoylation

Myristic acid may be linked via an amide bond to the a-amino group of the N-terminal glycine residue of selected proteins (Figure 9.18). The reaction is referred to as A -myristoylation and is catalyzed by myristoyl—CoAtprolein N-myris-toyltransferase, known simply as NMT. A -Myristoyl-anchored proteins include the catalytic subunit of cAMP-dependent protein kinase, the ppSff tyrosine kinase, the phosphatase known as calcineurin B, the a-subunit of G proteins (involved in GTP-dependent transmembrane signaling events), and the gag proteins of certain retroviruses, including the FHV-l virus that causes AIDS. 

A variety of cellular and viral proteins contain fatty acids covalently bound via ester linkages to the side chains of cysteine and sometimes to serine or threonine residues within a polypeptide chain (Figure 9.18). This type of fatty acyl chain linkage has a broader fatty acid specificity than A myristoylation. Myristate, palmitate, stearate, and oleate can all be esterified in this way, with the Cjg and Cjg chain lengths being most commonly found. Proteins anchored to membranes via fatty acyl thioesters include G-protein-coupled receptors, the surface glycoproteins of several viruses, and the transferrin receptor protein. 

TRAM was the fourth adapter discovered and has only been seen to have a role in TLR-4 signalling. It contains a TIR domain and a myristoylation site. When TRAM is myristoylated it becomes bound to the plasma membrane and can bind to TLR-4 through its TER. domain. TRAM then allows TRIF to bind it and activate the pathways associated with TRIF as outlined above for TLR-3. 

Veuillez F, Falson-Rieg F, Guy RH, Deshusses J, Buri P (2002) Permeation of a myristoylated dipeptide across the buccal mucosa Topological distribution and evaluation of tissue integrity. Int J Pharm 231 1-9... 

Fig. 3.13. Lipid anchors and basic regions as elements of the membrane association of proteins. Examples for proteins which exhibit basic residues near a lipid anchor, a) Src kinase (see ch. 8) possess a myristoyl anchor at the N-terminus as well as a stretch of basic residues, b) In Ki-Ras proteins (see ch. 9) there is a farnesyl residue at the C-terminus that serves as a lipid anchor, as weU as a stretch of Lys residues. Negatively charged head groups of phospholipids are shown as filled circles. X any amino acid.

A basic understanding of the structure of the BCR-ABLl chimeric protein is important to understanding how the multiple mutations that have been described in the ABLl kinase domain cause resistance. The c-ABLl protein is expressed in two splice forms that are known as la and lb (Fig. 1C). The la form is 19 residues shorter than lb. The lb form contains a myristoylation site on its second residue. The second residue is a glycine that appears to help regulate enzymatic activity, and its mutations to alanine prevents myristoylation and results in an activated kinase (50). The lb form also contains a cap region that is believed to stabilize the inactive eonfirmation of the kinase (51). The numbering system used to identify the amino acid residues where mutations occur is based on the shorter la form. 

Hantschel O, Nagar B, Guettler S et al. A myristoyl/phosphotyrosine switch regulates c-Abl. Cell 2003 112 845-857. 

There are two prominent types of mammalian cAMP-dependent protein kinases.51 61 The catalytic subunit is identical for both the 41-kDa peptide as isolated from beef heart has 350 residues and an N terminus blocked by a myristoyl (tetradecanoyl) group.62 One phosphoserine and one phosphothreonine are also present.51 The 50-kDa regulatory subunits vary in size and may also be subject to additional regulation by phosphorylation.63 Three-dimensional structures are known for both the catalytic62 64 65 and the regulatory66 subunits. A cyclic GMP (cGMP)-activated protein... 

EtP, ethanolamine phosphate) (b) a myristoylated protein (c) a prenylated protein (d) a palmitoylated protein. 

Raju, R. V., and Sharma, R. K. 1997. Demonstration and purification of a myristoyl-CoA binding protein from bovine cardiac muscle. Life Sci. 60 2145-2153. 

LpxM IpxM Acyltransferase Kdo2-penta-lipid A Myristoyl ACP Brozek and Raetz (1990)... 

Another wide application of mass spectrometry is the detection and characterization of post-translational modifications such as myristoylation, phosphorylation, disulfide bridging, etc. The detection and localization of post-transla-tional modifications has been a rapidly developing area of mass spectrometry due to the functional importance of these modifications in biological systems. An example of this was recently shown for the case of the human rhinovirus HRV14 [10]. Electron density maps from crystallography data indicated a myristoylation of VP4. Mass analysis of VP4 also indicated a mass difference of 212 Da (consistent with myristoylation of VP4). Additional experiments with proteolytic digestion and tandem mass spectrometry were able to localize the modification to the N-terminus of VP4. 

It is the structure of the major structural protein VPl that was best characterized (see below) (Liddington et al, 1991). The minor proteins VP2 and VPS share a G terminus, but VP2 has a myristoylated N-terminal addition of 100 residues. These proteins are mostly internal in the capsid, but the G terminus binds to VPl. 

Epand RE, Sayer BG, Epand RM. Induction of raft-hke domains by a myristoylated NAP-22 peptide and its Tyr mutant. FEBS J. 2005 272 1792-1803. 

The role of Arf in the in vivo activation of PLD remains unclear for Arfs 1-5, but there is more evidence for Arf6. Whereas Arfs 1-3 are associated with the Golgi, the subcellular distribution of Arfs 4 and 5 is unclear. In contrast, Arf6 is predominantly associated with the plasma membrane where it cycles between this membrane and the cytosol and a vesicular compartment as a function of its GTP/ GDP binding [105-109]. Stimulation of chromaffin cells induces translocation of Arf6 to the plasma membrane and an associated increase in PLD activity [109]. In addition, treatment of the cells with a myristoylated peptide corresponding to the N-terminal sequence of Arf6 inhibits PLD activation [109]. Similar results have been obtained wifh this peptide in myometrium [110]. 

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