Phosphatase specificity

Clarke, P. R., Hoffman, I., Draetta, G., and Karsenti, E. (1993). Dephosphorylation of cdc25-C by a type-2A protein phosphatase specific regulation during the cell cycle in Xenopus egg extracts. Mol. Biol. Cell 4 397—411. 

Primary cultured porcine or bovine brain capillary endothelial cells have been used as an in vitro model for the BBB. Recently, an immortalized cell line has been established from mouse, rat, and human brain capillary endothelial cells by infection with Simian virus 40 or transfection of SV40 large T antigen (45 -7). Tatsuta et al. established an immortalized mouse brain capillary endothelial cell line (MBEC4). The activity of y-glutamyl transpeptidase and alkaline phosphatase, specific marker enzymes for brain capillary endothelial cells, was half that in the brain capillary (45). Also, P-gp was expressed on the apical membrane of MBEC4 cells, which corresponds to the abluminal membrane of the brain... 

In general there are three phosphatase families alkaline, acid, and protein phosphatases. Alkaline phosphatases are typically dimers that contain three metal ions per subunit and have a pH optimum pH above 8. Acid phosphatases exhibit an optimum pH<7 and are usually divided into three classes low molecular weight acid phosphatases (<20 kDa), high molecular weight acid phosphatases (50-60 kDa), and purple acid phosphatases (which contain an Fe-Fe or Fe-Zn center at the active site). Phosphatases specific for I-l-P appear to be most similar (in kinetic characteristics but not in mechanism) to the alkaline phosphatases, but their structures define a superfamily that also includes inositol polyphosphate 1-phosphatase, fructose 1, 6-bisphosphatase, and Hal2. The members of this superfamily share a common structural core of 5 a-helices and 11 (3-strands. Many are Li+-sensitive (York et al., 1995), and more recent structures of archaeal IMPase proteins suggest the Li+ -sensitivity is related to the disposition of a flexible loop near the active site (Stieglitz et al., 2002). 

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. 

Bialojan, C., and Takai, A. 1988. Inhibitory effect of a marine-sponge toxin, okadaic acid, on protein phosphatases. Specificity and kinetics. Biochem J 256, 283-290. 

Phosphatases specific for such substrates as glucose-6-phosphate, fructose-l,6-bisphosphate, and phospho-glycolate help to drive metabolic cycles (Chapter 17). The 335-residue fructose-1,6-bisphosphatase associates to form a tetramer with D2 symmetry. ° The allosteric enzyme exists in two conformational states (see Chapter 11). Activity is dependent upon Mg + or other suitable divalent cation, e.g., Mn or Zn, and is further enhanced by K+ or NH3. While the dimetal sites depicted in Figs. 12-23 and 12-24 are quite rigid and undergo little change upon formation of complexes with substrates or products, the active site of fructose-1,6-bisphosphatase is more flexible. There are three metal-binding sites but they contain no histidine side chains and have been seen clearly only in a product complex. Perhaps because of the need for... 

Sadovsky, E., and Zuckerman, H., An alkaline phosphatase specific to normal pregnancy. Obstet. Gynecol. 26, 211-214 (1965). 

Houston B, Stewart AJ, Farquharson C (2004) PHOSPHOl-A novel phosphatase specifically expressed at sites of mineralisation in bone and cartilage. Bone 34 629-637... 

Interestingly, all these compounds are inactive against mouse lAPs, therefore, unsuitable for studies of lAP function in mouse models. The objectives of the current studies were the identification of small-molecule modulators of mouse lAP and, potentially, identification of novel selective scaffolds of human lAP. To this end, we applied and optimized the chemiluminescent assays that were successfully utilized for screening human lAP, TNAP, and FLAP and developed a novel assay for mouse lAP, analogous to the assays of other isozymes. These assays utilize CDP-star, a substrate of alkaline phosphatases specifically invented for and commonly utilized in blotting techniques [19, 20]. Development and utilization of the prototype plate-reader enzymatic assay for TNAP isozyme with CDP-star substrate was previously described in detail elsewhere [21]. 

Sents W, Ivanova E, Lambrecht C et al (2013) The biogenesis of active protein phosphatase 2A holoenzymes a tightly regulated process creating phosphatase specificity. FEBS J 280 644-661... 

Nucleoside diphosphates may be cleaved, as mentioned above, by inorganic pyrophosphatase, and a phosphatase specific for IDP, GDP, and UDP has also been described. Adenylate kinase also may convert ADP to adenylate (plus ATP). Studies of these enzymes have been reviewed by Keilley (4) and Morton (5). 

Gianinazzi-Pearson V, Gianinazzi S (1978) Enzymatic studies on the metabolism of vesicular arbuscular mycorrhiza n. Soluble alkaline phosphatase specific to mycorrhizal infection in onion roots. Physiol Plant Pathol 12 45-53... 

Meisler, M. H., Langan, T. A. Characterization of a phosphatase specific for phospho-rylated histone and protamine. J. biol. Chem. 244, 4961—4968 (1969). 

Nonspecific Phosphoesteroses. These enzymes cleave a variety of phosphoric acid derivatives, even synthetic substrates and lecithins (diesters of phosphoric acid with glycerol and choline). Such enzymes occur among other places in snake venom, and in the intestinal mucosa. Snake venom phosphatases specifically split 3 -phos-phate bonds, giving rise to 5 -monophosphates, whereas chemical hydrolysis yields a mixture of the 2 - and 3 -monophosphates. Free 3 -phosphate groups (the monoesters) are inhibitory in the enzymic reaction. 

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