Bacterial alkaline phosphatase activators

Several independent techniques have been used to identify bacterial alkaline phosphatase activity in natural bacterial assemblages. Phosphatase activity perse has often been detected using para-nitrophenyl phosphate as a substrate enzyme activity is expressed as rate of formation of the hydrolytic product, para-nitrophenol, which is easily detected spectrophotometrically because it absorbs light strongly at = 395 10 nm at pH > 9 (see Huber and Kidby, 1984, for a review of variations on this theme). Other substrates that have been used to detect alkaline phosphatase activity... 

Polakowski R, Craig D B, Skelley A and Dovichi N J 2000 Single molecules of highly purified bacterial alkaline phosphatase have identical activity J. Am. Chem. See. 122 4853-5... 

Metal ions also participate in the functioning of other nucleases, although the structural details of their participation are not nearly as established as those for staphylococcal nuclease. DNAse I also requires Ca for its catalytic activity.SI endonuclease, mung bean nuclease, and Physarum polycephalem nuclease require zinc ion either as cofactors or intrinsically for nuclease activity, and the restriction enzyme EcoRI may also require intrinsically bound zinc ion. In terms of how the zinc ion might function in these enzymes, one can look both to staphylococcal nuclease and to bacterial alkaline phosphatase for some... 

Bacterial alkaline phosphatase is the gene product of phoA, a member of the pho regu-lon (Table 9.1). When the pho regulon is induced by low external quantities of phosphate, synthesis of this alkaline phosphatase can represent as much as 6 mole% of total protein synthesis, and enzyme activity per cell can increase 1000-fold (Coleman and Gettins, 1983). The enzyme is synthesized as 43,000 Da monomers, which are transported to the periplasmic space and become active only after dimerization. As with many alkaline phosphatases, this enzyme accepts a broad range of substrates, which it hydrolyses at similar rates (Fernley and Walker, 1967 Reid and Wilson, 1971). Substrates are compounds with the general formula... 

Bacterial 5 -nucleotidase activity was first noted in marine systems as a means of turning over 5 -mononucleotides, conceivably released by phosphodiesterase or exonucleolytic activity (Ammerman and Azam, 1985). Although alkaline phosphatase can hydrolyse these substrates, it can be distinguished from 5 -nucleotidase activity by its sensitivity to micromolar concentrations of phosphate 5 -nucleotidase was virtually unaffected by 100 p,M phosphate, while 80% of alkaline phosphatase activity was lost under the same conditions (Ammerman and Azam, 1991a). Further, 5 -nucleotidase was competitively inhibited by 5 -guanidyl monophosphate, but almost completely... 

Stewart, A.J. and Wetzel, R.C. (1982b) Influence of dissolved humic materials on carbon assimilation and alkaline phosphatase activity in natural algal-bacterial assemblages. Freshwater Biology 12, 369-380. 

Methods based on chemiluminescent and bioluminescent labels are another area of nonisotopic immunoassays that continue to undergo active research. Most common approaches in this category are the competitive binding chemiluminescence immunoassays and the immunochemiluminometric assays. Chemiluminescence and heterogenous chemiluminescence immunoassays have been the subject of excellent reviews (91, 92). Detection in chemiluminescence immunoassays is based on either the direct monitoring of conjugated labels, such as luminol or acridinium ester, or the enzyme-mediated formation of luminescent products. Preparation of various derivatives of acridinium esters has been reported (93, 94), whereas a variety of enzyme labels including firefly or bacterial luciferase (70), horseradish peroxidase (86, 98), and alkaline phosphatase are commercially available. 

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