5600235565

I Konferencja Naukowa Enzymos Enzymy w nauce i przemyśle Wystąpienia Gości Honorowych_

Allosteric prostatic acid phosphatase: activity regulation, homotropic activators and heterotropic inhibitors

Prostatic acid phosphatase (PAP; EC 3.1.3.2) nonspecifically catalyzes hydrolysis of phosphomonoesters, as phospholipids, phosphosugars, phosphoproteins, nucleotides and nucleic acids. Its biological role in process of fertilization was not recognized yet. Initial reaction ratę vo was calculated from tangent of the kinetic curve (Excel), but catalytic constants (kcat, K0.5 and h) from Hill equation using Computer programs Ez-Fit and SigmaPlot.

v_0=(k_cat [EJ_0 ([S]_0 )^Ah)/((K_0.5 )^Ah+([S]_0)^Ah )    (1)

Enzyme-substrate saturation plots described by Hill eąuation (1) are sigmoidal. Hill cooperation coefficient h is higher than 1 (up to 4). Reaction's kinetics thus exhibits homotropic allostery (positive cooperativity). Substrates are homotropic activators of enzyme. Degree of cooperativity of PAP grows when enzyme concentration increases and depends on the Chemical character of substrate. Degree of cooperativity of PAP also grows when the water-pool radius of reverse micelle increases. Eąuilibrium of oligomeric PAP forms is suggested: monomer-dimer-tetramer, dependent on character of substrate (ligand) and on protein concentration in aąueous Solutions but on micellar dimensions. Hypothesis of ąuatemary structure changes was confirmed in physicochemical studies: by ultracentrifugation and fluorescence. MWC and KNF models do not describe allosteric character of PAP. Two known heterotropic inhibitors of allosteric PAP, tartrate and vanadate, were studied.

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