Alkaline phosphatase inhibition

Fig. 13. Representation of the formation of an EIS complex during alkaline phosphatase inhibition by L-phenylalanine. Compare with Fig. 9.

Fernley HN, Walker PG (1967) Studies on alkaline phosphatase. Inhibition by phosphate derivatives and the substrate specificity. BiochemJ 104 1011-1018... 

Reuter, Jr., J. G. (1983), Alkaline Phosphatase Inhibition by Copper Implications to Phosphorus Nutrition and Use as a Biochemical Marker of Toxicity, Limnol. Oceanogr. 28, 743-748. 

Y., Pekec, B., Jitchareon, ]., and Kalcher, K. (2013) Alkaline phosphatase inhibition-based amperometric biosensor for the detection of carbofuran. Int. 

Kim, I.Y., Han, S.Y., and Kang, T.S. et al. (2005). Pyrethroid insecticides, fenvalerate and permethrin, inhibit progesterone-induced alkaline phosphatase activity in T47D human breast cancer cells. Journal of Toxicology and Environmental Health—Part A—Current Issues 68, 2175-2186. 

Figure 6, Magnesium in concentrations of 0.001-10 mM/l. produces no significant activation of alkaline phosphatase in EAE, DEA, or 2A2M1P buffers. It activates the enzyme in carbonate buffer but inhibits it in glycine buffer.

Alkaline phosphatase assays based on 3-glycerophosphate now appears to be obsolete, and methods buffered by either glycine or barbital are also obsolete as these buffers inhibit ALP or are poor buffers. Serum alkaline phosphatase is known to be composed of several isoenzymes which presumably arise from bone, liver, intestine, and placenta. The placental alkaline phosphatase is known to be much more resistant to heat denaturation than the other isoenzymes, and this resistance provides a simple test for it (5). The other enzymes can be separated through the differential inhibition by phenylalanine, by electrophoresis and by specific antibodies. However, the clinical usefulness of the results obtained is in doubt (23). 

Yamaguchi and Gao, 1998 Rat femoral-metaphyseal tissues cultured for 48 h with bone resorbing factors PTH, PGE2 or EPS) +/- genistein measured bone calcium content, acid and alkaline phosphatases Genistein (10 Yi O M) inhibited bone resorption. Effect reversed by anti-estrogen, tamoxifen. 

Alkaline phosphatases [AP, orthophosphoric-monoester phosphorylase (alkaline optimum) EC 3.1.3.1] represent a large family of almost ubiquitous isoenzymes found in organisms from bacteria to animals. In mammals, there are two forms of AP, one form present in a variety of tissues and another form found only in the intestines. They share common attributes in that the phosphatase activity is optimal at pH 8-10, is activated by the presence of divalent cations, and is inhibited by cysteine, cyanides, arsenate, various metal chelators, and phosphate ions. Most conjugates created with AP utilize the form isolated from calf intestine. 

M. Ayyagari, S. Kametkar, R. Pande, K.A. Marx, J. Kumar, S.K. Tripathy, J. Akkara, and D.L. Kaplan, Chemiluminescence-based inhibition kinetics of alkaline phosphatase in the development of a pesticide biosensor. Biotechnol. Prog. 11, 699-703 (1995). 

Inhibited calcium accumulations in bone and increased alkaline phosphatase activity of medium (Kaji etal. 1988). 

Alkaline phosphatase is an enzyme represented by various isoforms in many tissues such as liver, bone, intestine, placenta, some tumors and in leukocytes. Addition of 1 mM levamisole to the chromogen/substrate will inhibit endogenous alkaline phosphatase activity, with the exception of the intestinal isoform. If necessary, this can be blocked with a weak acid wash, such as 0.03 0.5 N HC1 or 1 M citric acid. 

Not all inhibitors fall into either of these two classes but some show much more complex effects. An uncompetitive inhibitor is defined as one that results in a parallel decrease in the maximum velocity and the Km value (Figure 8.8). The basic mode of action of such an inhibitor is to bind only to the enzyme-substrate complex and not to the free enzyme and so it reduces the rate of formation of products. Alkaline phosphatase (EC 3.1.3.1) extracted from rat intestine is inhibited by L-phenylalanine in such a manner. 

Inhibits serine proteases such as trypsin and chymotrypsin. Also inhibits cysteine proteases (reversible by reduced thiols) and mammalian acetylcholinesterase Inhibits ATPase, alkaline phosphatase and tyrosine phosphatase Reagent for maintaining -SH groups in the reduced state. Effective for reducing protein disulfide bonds prior to SDS-PAGE... 

Because alkaline phosphatase converts ATP or GTP to their respective nucleosides, use of alkaline phosphatase to deplete ATP or GTP should be reserved for those cases where this does not present a problem Furthermore, alkaline phosphatase is potently inhibited by orthophosphate so higher than would be anticipated amounts of enzyme are required for efficiently depleting ATP or GTP. 

Enzymes activities are particularly sensitive to the anticoagulant used in collecting the specimen. Heparin inhibits acid phosphatase (W16) and muramidase (Z5). Amylase activity is inhibited by oxalate or citrate (MIO), and lactic dehydrogenase and acid phosphatase lose activity in oxalate (C2). Alkaline phosphatase is stable in oxalate, oxalate-fluoride, or heparin, but 25 mAf citrate inhibits 50% of the activity, and as little as 50 mlf EDTA is completely inhibitory (B19). Leucine aminopeptidase is inhibited by EDTA, as is creatine phosphokinase (F3). Amylase activity has been reported to be only 83% of that in serum when oxalate or citrate-plasma is used (MIO). Heparin plasma appears to have no inhibitory effect. Despite the fact that clotting factor V is not stable in oxalate or EDTA, these are often used as anticoagulants to obtain plasma for prothrombin determinations (Z2, Z4). 

Liver injury is clinically defined as an increase of serum alanine amino transferase (ALT) levels of more than three times the upper limit of normal and a total bilirubin level of more than twice the upper limit of normal [4]. The clinical patterns of liver injury can be characterized as hepatocellular (with a predominant initial elevation of ALT), cholestatic (with an initial elevation of alkaline phosphatase) or mixed. The mechanisms of drug-induced hepatotoxicity include excessive generation of reactive metabolites, mitochondrial dysfunction, oxidative stress and inhibition of bile salt efflux protein [5]. Better understandings of these mechanisms in the past decades led to the development of assays and models suitable for studying such toxic mechanisms and for selecting better leads in the drug discovery stage. 

Currently, only a handful of examples of unique protein carboxylate-zinc interactions are available in the Brookhaven Protein Data Bank. Each of these entries, however, displays syn coordination stereochemistry, and two are bidentate (Christianson and Alexander, 1989) (Fig. 5). Other protein structures have been reported with iyw-oriented car-boxylate-zinc interactions, but full coordinate sets are not yet available [e.g., DNA polymerase (Ollis etal., 1985) and alkaline phosphatase (Kim and Wyckoff, 1989)]. A survey of all protein-metal ion interactions reveals that jyw-carboxylate—metal ion stereochemistry is preferred (Chakrabarti, 1990a). It is been suggested that potent zinc enzyme inhibition arises from syn-oriented interactions between inhibitor carboxylates and active-site zinc ions (Christianson and Lipscomb, 1988a see also Monzingo and Matthews, 1984), and the structures of such interactions may sample the reaction coordinate for enzymatic catalysis in certain systems (Christianson and Lipscomb, 1987). 

The type of inhibition of chicken intestine alkaline phosphatase by forphenicine was not competitive, but uncompetitive with the stibstrate. Its derivative, forphenicinol, which contains a hydroxymethyl grorp instead of the formyl group in the forphenicine molecule, did not inhibit alkaline phosphatase but, it did bind to cells. Forphenicinol enhanced delayed-type lOT>ersensiti-vity (13,14) and the phagocytic activity of macrophages. 

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