Protein tyrosine phosphatase Mechanism

Denu, J. M. and Dixon, J. E. (1998) Protein tyrosine phosphatases mechanisms of catalysis and regulation. Curr. Opin. Chem. Biol. 2, 633-641. 

Insulin binding to the extracellular side of cell membranes initiates the insulin cascade , a series of phosphorylation/dephosphorylation steps. A postulated mechanism for vanadium is substitution of vanadate for phosphate in the transition state structure of protein tyrosine phosphatases (PTP).267,268 In normal physiological conditions, the attainable oxidation states of vanadium are V111, Viv and Vv. Relevant species in solution are vanadate, (a mixture of HV042-/ H2VOO and vanadyl V02+. Vanadyl is not a strong inhibitor of PTPs, suggesting other potential mechanisms for insulin mimesis for this cation. 

Majeti, R. and Weiss, A. (2001). Regulatory mechanisms for receptor protein tyrosine phosphatases. Chem. Rev. 101, 2441-2448. 

The catalytic center of the protein tyrosine phosphatases includes ca. 230 amino acids and contains the conserved sequence motif HA -C-(X)5-R-S/T-G/A/P (X is any amino acid) which is involved in phosphate binding and in catalysis and is part of a loop known as the P loop. The available structural data on the catalytic domains of protein tyrosine phosphatases indicate that the mechanism shown schematically in Fig. 8.17 is likely (see Tainer and Russell, 1994). The invariant Cys and Arg residues of the P loop have a central function in binding and cleavage of the phosphate residue. 

The fact that cellular activity of protein tyrosine phosphatases by far exceeds that of protein tyrosine kinases suggests that there is strict control of the dephosphorylation rate in a cell. The mechanisms are those already highlighted in previous chapters as central elements of regulation of activity of signal molecules. 

Another mechanism of regulation of protein tyrosine phosphatases is via Ser/Thr phosphorylation. Specific phosphorylation of protein tyrosine phosphatases by Ser/ Thr-specific protein kinases of types A and C has been reported (see Neel and Tonks, 1997). This observation indicates the possibility that signal transductions via Ser/Thr kinases and via Tyr kinases/phosphatases may cooperate and that different signal pathways may be crosslinked in this way. 

Down-regulation of T-cell protein tyrosine phosphatase in IM-resistant cells may represent a novel mechanism for IM resistance (40). A recently published analysis of data from the IRIS trial demonstrated that trough blood levels of IM and its active major metabolite, CGP74588, correlated with achievement of CCR and MMR (41). 

The protein tyrosine phosphatases also exist as several families with numerous functions in control of transcription, growth, differentiation, and metabolism.741-743 These enzymes function by a doubledisplacement mechanism, as in Eq. 12-38, but with a cysteine side chain rather than serine. The cysteine is present in the conserved sequence (H/V)CX5R(S/T). The arginine binds the phospho group and helps to stabilize the transition state, which probably is metaphosphate-like.742... 

Tjernberg, A., et al., Mechanism of action of pyridazine analogues on protein tyrosine phosphatase IB (PTP1B). Bioorg Med Chem Lett, 2004, 14, 891-895. 

Huyer, G., S. Liu, J. Kelly, J. Moffat, P. Payette, B. Kennedy, G. Tsaprailis, M.J. Gresser, and C. Ramachandran. 1997. Mechanism of inhibition of protein-tyrosine phosphatases by vanadate and pervanadate. J. Biol. Chem. 272 843-851. 

Studies of the oxidation of organic sulfides with amino acid-derived ligands in acetonitrile revealed very little difference between the mechanism of their oxidation and that of halides, except for one major exception. Despite the fact that acid conditions are still required for the catalytic cycle, hydroxide or an equivalent is not produced in the catalytic cycle, so no proton is consumed [48], As a consequence, there is no requirement for maintenance of acid levels during a catalyzed reaction. Peroxo complexes of vanadium are well known to be potent insulin-mimetic compounds [49,50], Their efficacy arises, at least in part, from an oxidative mechanism that enhances insulin receptor activity, and possibly the activity of other protein tyrosine kinases activity [51]. With peroxovanadates, this is an irreversible function. Apparently, there is no direct effect on the function of the kinase, but rather there is inhibition of protein tyrosine phosphatase activity. The phosphatase regulates kinase activity by dephosphorylating the kinase. Oxidation of an active site thiol in the phosphatase prevents this down-regulation of kinase activity. Presumably, this sulfide oxidation proceeds by the process outlined above. 

Huyer, G. 1997. Mechanism of inhibition of protein tyrosine phosphatases by vanadate and pervanadate. J. Biolog. Chem. 272 843-851. 

Fig. 2. Mechanisms of VO(OPT)-enhanced neurogenesis after brain ischemia in the hippocampal dentate gyrus. VO(OPT) inhibits protein tyrosine phosphatase IB, thereby stimulating Akt and ERK signaling. The downstream targets of Akt and ERK include HIF-la, GSK-3/3, and CREB, which are involved in the proliferation, migration, and maturation of the neural progenitor cells.

Liu S, Zhou B, Yang H et al (2008) Aryl vinyl sulfonates and sulfones as active site-directed and mechanism-based probes for protein tyrosine phosphatases. J Am Chem Soc 130 8251-8260... 

Another AT2 pathway linked to apoptosis leads to a stimulation of a soluble protein tyrosine phosphatase, SHP-1, an enzyme that associates with insulin receptor substrate (IRS)-2 (Cui et al. 2002). Overexpression of a dominant negative form of SHP-1 in PC12W cells has been found to attenuate AT2 receptor-mediated inhibition of insulin signaling. Since insulin activates Akt, it is interesting that the mechanism of apoptosis in the angiotensin II-treated PC12W cell involves the dephosphorylation and inactivation of Akt. In NIE-115 neuroblastoma cells,... 

Reversible chemical modification of enzymes, which was discovered in 1955 by Edmond Fischer and Edwin Krebs [58], is a more prevalent mechanism for cellular signaling switching. Fischer and Krebs showed that enzymes can be turned from an inactive form to an active form via phosphorylation of certain residues of the protein. Enzymes that catalyze phosphorylation (addition of a phosphate group coupled with ATP or GTP hydrolysis) are called protein kinases. Enzymes that catalyze dephosphorylation (which is not the reverse reaction of the phosphorylation) are called phosphatases. For example, a protein tyrosine phosphatase is an enzyme that catalyzes the removal of a phosphate group from a tyrosine residue in a phosphorylated protein [57],... 

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). 

Figure4.1 Protein tyrosine phosphatase IB (2hnq (a)) and CDC25B (lcwt (b)) employ the same catalytic mechanism for hydrolysis of phosphorylated substrates but share no sequence homology and exhibit very different folds. The similarity of their binding sites can, however, be detected on the level of the interaction properties exposed to a ligand.

Burke, T. R. and Zhang, Z. Y. (1998) Protein-tyrosine phosphatases Structure, mechanism, and inhibitor discovery. Biopolymers 47, 225-241. 

Many members of this class of receptors have an enzymatic activity known as a protein tyrosine kinase within their cytoplasmic segment. This kinase phosphorylates tyrosine residues in the receptors themselves (autophosphory lation), and in other proteins to initiate biochemical cascades. Phosphorylatipn of tyrosine can be reversed by protein tyrosine phosphatases, which are also present in all cells (Shenolikar and Naim, 1990). Tyrosine phosphatases form a diverse family of proteins, some of which are cytosolic while others are transmembrane molecules analogous to receptors. Some members of the transmembrane class may be involved in the mechanism of bacterial and viral infections (Tonks, 1991). Thus, kinases and phosphatases together act as on-off switches in the a ctivation of receptors and other proteins. 

Phosphatases hydrolyze phosphate groups from phosphopro-teins, thereby reversing the action of protein kinases. Two classes of protein phosphatases exist with distinct structures, substrate specificities, and mechanisms protein serine/threonine phosphatases and protein tyrosine phosphatases. A few dualspecificity phosphatases also are known that are similar in structure to protein tyrosine phosphatases. 

Figure 5 (a) Proposed catalytic mechanism of protein tyrosine phosphatases, (b) Structure of PTPl B (gray) with a phosphopeptide substrate (goid) showing the relative positions of catalytic residues to the phosphotyrosine moiety. 

Kolmodin K, Aqvist J. The catalytic mechanism of protein tyrosine phosphatases revisited. EEBS Lett. 2001 498 208-213. 

Signaling. Protein tyrosine phosphatases, such as the molecule CD45 expressed in both B cells and T cells, play important roles in activating such protein tyrosine kinases as Fyn and Lck, which are quite similar to Src. Suggest a mechanism for the activation of such protein kinases by the removal of a phosphate from a phosphotyrosine residue. 

---