Tyrosine-specific phosphatases

Fig. 3.8 The protein tyrosine phosphatases (PTPs) with the C(X)gR motif are divided into tyrosine-specific PTPs the VHl-like dual specificity, serine/threonine- and tyrosine-specific phosphatases the CDC25 phosphatase and the low molecular weight (LMW) phosphatases. The low molecular weight phosphatases are acid phosphatases without distinct regulatory or targeting domains. Their function is not known. The tyosiiie-speoifio phosphatases are further subdivided into receptor-like and non-reoeptor-like phosphatases. (This scheme is reproduced with permission of the authors and Trends Biochem. Sci. from ref. 76.)...

Fig. 3.9 (a) A ribbon presentation of the structure of the human tyrosine-specific phosphatase IB, (PTPIB). It is a single-domain protein with a seven-stranded, mixed b-sheet, flanked by a-helices. The catalytic site is in the central region, in a shallow cleft. The phosphate-recognition site is characteristic for PTPs. It is formed by a loop containing the C(X5>R motif, which contains the catalytically essential cysteine and arginine residues. (Reproduced with permission of the authors, D.Barford, A. J. Flint, and N. K.Tonks, and Science from ref. 78.)... 

Tyrosine PPs are classified in four families, depending on their biological function and aminoacid sequence tyrosine-specific phosphatases (TSPs), vaccinia virus HI (VHl)-like phosphatases, cdc25 phosphatases... 

Receptor-like protein tyrosine phosphatases Receptor tyrosine kinases Stress-activated PK/cJun N-terminal kinase SAPK/ERK kinase 1 Src homology domain 2 Transforming growth factor-(3 Tumour necrosis factor 12-O-tetradecanoylphorbol-13-acetate Tyrosine specific phosphatases... 

It is not clear whether V(V) or V(IV) (or both) is the active insulin-mimetic redox state of vanadium. In the body, endogenous reducing agents such as glutathione and ascorbic acid may inhibit the oxidation of V(IV). The mechanism of action of insulin mimetics is unclear. Insulin receptors are membrane-spanning tyrosine-specific protein kinases activated by insulin on the extracellular side to catalyze intracellular protein tyrosine phosphorylation. Vanadates can act as phosphate analogs, and there is evidence for potent inhibition of phosphotyrosine phosphatases (526). Peroxovanadate complexes, for example, can induce autophosphorylation at tyrosine residues and inhibit the insulin-receptor-associated phosphotyrosine phosphatase, and these in turn activate insulin-receptor kinase. 

The extent of tyrosine phosphorylation of signal proteins is determined both by the activity of the tyrosine kinases and also the activity of tyrosine-specific protein phosphatases. If the total activity of both enzymes in the cell is considered, it is found that there is a preponderance of protein tyrosine phosphatase activity compared to tyrosine kinase activity. In contrast, the activities of the Ser/Thr-specific protein kinases and protein phosphatases are approximately balanced. It is estimated that the activity of the protein tyrosine phosphatases is about 3-4 orders of magnitude higher than the activity of the protein tyrosine kinases. With this relationship between the activities, it is not surprising that the net level of tyrosine phosphorylation in the cell is very low and that tyrosine phosphorylation is often only transient. Consequently, it took a relatively long time until the importance of tyrosine phosphorylation for signal transduction was assessed correctly. 

True enough, treatment of PAP with FMPP resulted in a time-dependent inactivation of the enzyme. Competitive inhibitors of PAP protected against inactivation. The authors suggest that FMPP represents a useful basic structure which can be incorporated into the design of more specific phosphatase inhibitors for example, the modified tyrosine 77 could be incorporated into a particular peptide to give a suicide substrate that is selective for a protein phosphatase which preferentially hydrolyses that peptide. 

The phosphorylated receptor appears to be capable of being dephosphorylated by the action of endogenous cytosolic phosphatases, as incubation of cells with vanadate, which inhibits the action of tyrosine-phosphate-specific phosphatases, increased the phosphorylation state of the receptor [60,61]. The significance of this observation has yet to be ascertained. 

On the other hand, a particular protein function can be realized with different protein folds, and an example of this are protein phosphatases. Protein phosphatases feature two distinctively different catalytic mechanisms for hydrolytically cleaving phosphorylated amino acid residues. The active sites of serine/threonine protein phosphatases (PPs) contain two metal centers that directly activate a water molecule for nucleophilic attack of the phosphate ester bond. In contrast, protein tyrosine phosphatases (PTPs) [105] possess a Cys residue present in the active site loop containing the conserved PTP signature motif HCXXXXXRS. The Cys sidechain acts as the attacking nucleophile in the formation of a phosphocysteine intermediate, which is eventually hydrolyzed by a water molecule [106], The same catalytic mechanism is also shared by dual-specificity phosphatases (see below). 

For dephosphorylation reactions, many enzymes with different specificities are available There are phosphatases, specific for phosphotyrosyl residues, for Ser/Thr-bound phosphates, and dual-specificity phosphatases, recognizing both phosphotyrosyls and phosphoserines Tyrosine phosphatases and dual-specificity phosphatases have already been introduced (Chapter 3). Here, the properties of serine/threonine phosphatases will be described and their regulation by cellular relocation introduced. Much of what we know about the regulation of this class of phosphatases we owe to the work of P. Cohen and his colleagues. Table 7.1 lists common phosphoserine/phosphothreonine phosphatases of eukaryotes. 

The mode of phosphorylation can also be determined enzymatically using phosphatases that are specific for either the serine/threonine or the tyrosine linkages. Also, tyrosine specific phosphorylation can be detected using phosphotyrosine specific antibodies (Kamps and Sefton 1988). 

Key words Tyrosine phosphorylation. Phosphatases, PTP, Structure, Fimction, Substrate recognition, Specificity, Human disease, PTPIB, SHP2, Inhibitors... 

Pulido B., Hooft van Huijsduijnen R (2008) Protein tyrosine phosphatases dual-specificity phosphatases in health and disease. FEBS J 275 848-866... 

Arrest in Gx phase can now be achieved in at least two ways, depending on the substrates of the Chkl and Chk2 enzymes. In one rapid way, the dual specificity phosphatase Cdc25C is phosphorylated on Ser 123 and is thereby targeted for ubiquitina-tion and degradation in the proteasome pathway. The lack of this enzyme locks the CDK2 kinase in the inactive form phosphorylated on threonine 14 and tyrosine 15.The cyclin E-CDK2 complex that is required for entry into S phase is inhibited, and the cell cycle arrests at Gj/S. It should be noted that the scheme in Fig. 13.19 is only a minimal scheme that does not address the participation of numerous other proteins that function as adaptors or structural proteins in these processes. 

The PTPs catalyze the hydrolysis of phosphorylated tyrosine residues in proteins, to yield the free tyrosine side chains and inorganic phosphate. They are classified according to substrate specificity (1) tyrosine-specific PTPs, such as the Yersinia PTP (YopH) and the mammalian PTPIB and PTPl, which in vivo hydrolyze only pTyr residues as well as (2) the dual-specificity phosphatases (DSPs), such as the human VHR and Cdc25, which hydrolyze pTyr and pSer and pThr residues of protein substrates. Based on their cellular localization, PTPs are classified as receptor-like or intracellular. ... 

Protein tyrosine phosphatases. Phosphoprotein phosphatases are integral components of the signahng systems operated by protein kinases (Sun and Tonks, 1994). Cloning data show the protein tyrosine phosphatases (PTPs) to be a family of multidomain proteins having exceptional diversity. They can be broadly divided into two groups, the transmembrane or receptor-like PTPs and the cytosolic PTPs. None of these are related to the serine-threonine specific phosphatases. This is in contrast to the protein kinases (Seer-Thr and Tyr specific), which share a common ancestry. Unlike the Ser-Thr phosphatases, in which substrate specificity is determined by associated targeting subunits, the Tyr phosphatases are all monomeric enzymes. 

The cytosolic PTPs are also classified according to their domain structures, which are understood to act as localization signals, directing the enzymes to the nucleus or cytoskeleton. Important subclasses, SHP-1 and SHP-2, possess SH2 domains. Others are characterized by the presence of PEST sequences (Pro-Glu/Asp-Sere/Thr) in the vicinity of the C-temtinus. Another subclass comprises dual specific phosphatases, which can dephosphorylate at both Tyr and Ser/Thr residues. The dual specific phosphatases also have homology with Cdc25, a regulator of mitosis in Schizoscccharomyces pombe. This activates cyclin dependent kinase-2 by dephosphorylation of adjacent threonine and tyrosine residues. 

PTPs are classified on the basis of their structure and sequence [3], They can be subgrouped into classical nonreceptor-type nontransmembrane PTPs, receptorlike membrane-localized PTPs, and the dual-specificity phosphatases (DUSPs) [4], Classical PTPs mostly recognize phospho-tyrosine as their substrate, whereas DUSPs are also known to dephosphorylate proteins on serine or threonine, and even phospholipids or RNA can be substrates [3], All PTPs contain a conserved CXXXXXR motive (single-letter amino acid code). The cysteine is the catalytically active moiety. Another residue that is involved in the catalytic mechanism (with some exceptions [5]) is an aspartic acid in the so-called WPD (Trp-Pro-Asp) loop, which is distinct from the active site [4] (Figure 1). 

Some phosphatases are nonspecific, catalyzing the hydrolysis of a wide variety of substrates. Other phosphatases are small-molecule specific and hydrolyze a particular small-molecule phosphate ester or structurally similar substrates. Phosphoprotein-specific phosphatases accept particular phosphorylated proteins or peptides as substrates. The phosphoprotein-specific phosphatases can be subdivided into subgroups those specific for proteins phosphorylated on tyrosine those specific for proteins phosphorylated on serine or threonine and the so-called dual-specific enzymes, which accept both classes of phosphorylated proteins. More recently, a group of protein histidine phosphatases has been described. 

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