Protein phosphoiylation

MENKE, F.L., PARCHMANN, S., MUELLER, M.J., KUNE, J.W., MEMELINK, J., Involvement of the octadecanoid pathway and protein phosphoiylation in fungal elicitor-induced expression of terpenoid indole alkaloid biosynthetic genes in Catharanthus roseus. Plant Physiol., 1999,119, 1289-1296. 

More than 60 different proteins were identified, and several phosphoiylated peptides were characterized using two different strategies (Toda et al, 2000). The first one was the use of affinity separation of proteins phosphoiylated on tyrosine after iimnunoprecipitahon, the second one was metal chelate affinity chromatography on ferric ions. 

Another global neuroproteomic study carried out by Collins et al, (unpublished results) focused on the neurophosphoproteome, i.e. the collection of phosphoproteins at the synapse. Protein phosphoiylation is an essential regulator of protein function and signaling mechanisms in any biological system. It has been estimated that one third of all proteins are phosphorylated in the mammalian cell and if it is assumed that one third of all proteins are expressed in the brain, then a conservative estimate of the number of phosphoproteins expressed in the brain would be in the order of... 

Certain serine, tyrosine or threonine residues in proteins can be phosphorylated by protein kinases and dephosphorylated by protein phosphatases (e.g. Chapters 25 and 27). This causes conformational changes in enzyme proteins which increase or decrease activity. This very important regulatory mechanism is confirmed since protein kinases and phosphatases amount to 5% of the proteins encoded by the human genome. Current research has shown that abnormalities of protein phosphoiylation are associated with diseases such as cancer, diabetes and inflammation, the protein kinases and phosphatases are therefore targets for future drug therapy... 

Boss, W.F., Yang, W., Tan, Z. and Cho, M. Regulation of phosphatidyl-inositol-4-kinase by protein phosphoiylation a plasma membrane-cytoskeletal connection, in P. Mazliak and J.C. Kader (eds.). Plant Lipid Metabolism, Kluwer Academic Publishers, Dordrecht, 1995, pp. 219-223. 

A growing number of protein crystal structures has provided solid evidence that in many phosphoesterase enzymes, two and sometimes even three, di- or trivalent metal ions are involved in substrate transformation. Consequently, the high catalytic efficiency is, in part, the result of a perfectly coordinated catalytic cooperation of the metal ions. Dinu-clear phosphoiyl transfer enzymes have been discussed thoroughly in recent reviews [1-3]. Therefore, this chapter (Section 2) only gives a brief description of enzymes for which two-metal promotion of phos-phoester hydrolysis was proposed on the basis of detailed mechanistic or crystallographic studies (Table 1). 

Figure 14.10 Diagrammatic representation of regulation of the opening of an ion channel by phosphoiylation of a protein in the channel. The neurotransmitter-receptor complex functions as a nucleotide exchange factor to activate a G-protein which then activates a protein kinase. This is identical to control of G-proteins in the action of hormones (Chapter 12, see Figure 12.21). Phosphorylation of a protein in the ion channel opens it to allow movement of Na+ ions. The formation of the complex, activation of the G-protein and the kinase takes place on the postsynaptic membrane. An example of the structural organisation and the involvement of a G-protein is shown in Chapter 12 (Figure 12.6).

Eugene Kennedy and Albert Lehninger showed in 1948 that, in eulcaiyotes, the entire set of reactions of the citric acid cycle takes place in mitochondria. Isolated mitochondria were found to contain not only all the enzymes and coenzymes required for the citric acid cycle, but also all the enzymes and proteins necessaiy for the last stage of respiration—electron transfer and ATP synthesis by oxidative phosphoiylation. As we shall see in later chapters, mitochondria also contain the enzymes for the oxidation of fatty acids and some amino acids to acetyl-CoA, and the oxidative degradation of other amino acids to a-ketoglutarate, succinyl-CoA, or oxaloacetate. Thus, in nonphotosynthetic eulcaiyotes, the mitochondrion is the site of most energy-yielding... 

Assays based on the ability of these toxins to inhibit protein phosphatases are a good example of this and are particularly usefirl since they can provide an indication of the biochemical activity of these toxins. These assays are rapid and sensitive and involve the measuring of the inhibitory effect of microcystins on the release of phosphate from phosphoiylated protein substrates (Bell 1994). 

Caution should however be taken in determining the proteins that are specifically tyrosine phosphoiylated in response to various agonists. This can be illustrated with thrombin, where studies have shown that removal of the autocrine stimulation in platelets during activation strongly reduces the tyrosine phosphorylation stimulated by thrombin (Figure 9.4.9.2). 

Borsch-HauboM AG, Kramer RM, and Watson SP. (1995). Cytosolic phospholipase A2 is phosphoiylated in collagen- and thrombin-stimulated human platelets independent of protein kinase C and mitogen-activated protein kinase. J. Biol. Chem. 270,25885-25892. 

After the insulin receptor catalyzes its own phosphorylation, the receptor is activated in a way that provokes it to phosphoiylate other proteins. The main substrate is a cytoplasmic protein called IRS. IKS stands for insulin responsive substrate. IRS has a molecular weight of 131,00(1 (131 kDa). The phosphorylation of IRS is represented by (Mothe and Obberghen, 1996) ... 

---