Acid phosphatase, prostatic

The acid phosphatases include all phosphatases that hydrolyze phosphate esters with an optimum pH of less than 7.0. They are present in the lysozymes of the secretory epithelial cells. Although acid phosphatase is produced primarily by the prostate gland, it is also found in erythrocytes, platelets, leukocytes, bone marrow, bone, liver, spleen, kidney, and intestine. 

The clinical use of PAP has been replaced by PSA. PAP is not as sensitive as PSA for screening or for detection of early cancer. The clinical use of PAP is restricted to confirmation of metastatic prostate cancer and staging of prostate cancer. It is less likely to be elevated in BPH than is PSA. Currently the method of choice for PAP is the measurement of its enzymatic activity. 

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Babson proposed a-naphthyl phosphate as an essentially specific substrate for the activity of prostatic acid phosphatase in serum (104). However Marshall, Price, and Amador found that this substrate is not specific for the prostatic enzyme because urine of human females contain 50 times more acid a-naphthyl phosphatase than male serum and 50% as much activity as male urine. Platelets have significant activity and the serum activity can increase to abnormal values following clotting. These workers also observed elevated activities in females with skeletal metastases of the breast. In 50 hospitalized male patients who had no evidence of prostatic cancer and 25 hospitalized female patients, the incidence of false positive results was 12%, a magnitude sufficient to preclude meaningful clinical interpretation (105). 

Ying, S. H. and Day, E. Serim prostatic acid phosphatase levels in proved cases of carcinoma or benign hypertrophy of the prostate. Cancer (1956), , 228-233. 

Very recently, a sandwich assay for prostatic acid phosphatase antigen was carried out using two cascaded enzyme reactions to provide amplification of the immunochemical event. In one format, an optical readout was used whereby a forma-zan dye was generated by reaction of a dye precursor and NADH generated from the second enzyme cycle. In the electrochemical format, the NADH generated in the second enzyme cycle was used to reduce Fe(CN) to FeCCN) " which was then detected amperometrically. While the use of Fe(CN) in ECIA has appeared in the... 

Note PCa, prostate cancer WM, white male DES, diethylstilbestrol AR, androgen receptor PSA, prostate specific antigen and PAP, prostatic acid phosphatase. 

Lin M-F, Meng T-C, Rao PS, et al. Expression of human prostatic acid phosphatase correlates with androgen-stimulated cell proliferation in prostate cancer cell lines. J. Biol. Chem. 1998 273 5939-5947. 

Provenge is a cancer vaccine using cell therapy technique. Dendritic cells are removed from patients. These cells are treated with the prostate-specific antigen prostatic acid phosphatase (PAP), which is present in 95% of prostate cancer cases. The activated dendritic cells are returned to the patients and they stimulate the T cells to destroy cancer cells expressing the PAP, thus treating the tumor. 

Prostate adenocarcinomas Prostate specific antigen (PSA) (25) Prostatic acid phosphatase (PAP)... 

AGIRE computer program for, 249, 79-81, 225-226 comparison to analysis based on rates, 249, 61-63 complex reactions, 249, 75-78 experimental design, 249, 84-85 inhibitor effects, 249, 71-75 potato acid phosphatase product inhibition, 249, 73-74 preliminary fitting, 249, 82-84 prephenate dehydratase product inhibition, 249, 72-73 product inhibition effects, 249, 72-73 prostate acid phosphatase phenyl phosphate hydrolysis, 249, 70 reactions with two substrates, 249, 75-77 reversible reactions, 249, 77-78 with simple Michaelian enzyme, 249, 63-71 [fitting equations, 249, 63] with slow-binding inhibitors, 249, 88 with unstable enzymes, for kinetic characterization, 249, 85-89. 

Although detailed structural as well as mechanistic knowledge of an enzyme is desirable, it is by no means necessary in order to design a suicide substrate. This has been shown by Myers and Widlanski (1993) who have designed a simple inhibitor of human prostatic acid phosphatase (PAP), an enzyme that is believed to be involved in the regulation of androgen receptor activity in prostate cells. Since the enzyme shows a preference for hydrolysis of aryl phosphates, the 4-(fluoromethyl)-phenyl phosphate (FMPP) was prepared as a substrate that would, on hydrolysis by the... 

Table 7.8 Hydrolysis of phosphate esters by prostatic acid phosphatase at pll 5 and 37"C" ...

Group Substrate Normal range (units/100 ml) Serum plus prostatic acid phosphatase Serum plus erythrocytic acid phosphatase Heated serum Relative specificity for prostatic acid phosphatase... 

Most investigators utilize p-nitrophenyl or a-naphthyl phosphate as substrate. The determination of serum prostatic acid phosphatase was developed by Fishman and Lemer (34) based on the d-(+)-tartrate inhibition of prostatic enzyme discussed below. Babson et al. (35, 36) demonstrated that a-naphthyl phosphate was much more easily split by prostatic than red cell phosphatase. Table V (35) shows the results obtained when prostatic or red cell phosphatase was added to human serum which had been incubated at pH 8.6 for 1 hr at 37° to destroy all endogeneous phosphatase activity. The table shows the superiority of a-naphthyl phosphate as substrate. 

A spectrofluorometric method for the estimation of acid phosphatase has been devised. It uses a-naphthyl phosphate as substrate thus, it is somewhat more specific for prostatic acid phosphatase than most (37). 

Fig. 1. Prostatic acid phosphatase activity as a function of pH ( ) phenyl phosphate (O) p-nitrophenyl phosphate and (A) /8-glycerophosphate. Buffers Ac, acetate Cit, citrate and tris. From Nigam et al. (88).

Reiner and his colleagues (40) demonstrated that fluoride inhibition of prostatic acid phosphatase showed interesting and unexpectedly complex kinetics. The unusual nature of the inhibition can readily be appreciated from Fig. 4 (40). As the fluoride concentration is increased over a 1000-fold range, the extent of inhibition rises and then subsequently falls with a further increase of inhibitor. At lower fluoride concentration, the inhibition is clearly competitive. Two possibilities were explored for an explanation of these unusual concentration effects of inhibition. There could be two forms of fluoride in the reaction mixtures the inhibitory form and the second which predominates at higher... 

Fig. 3. Surface inactivation rate of prostatic acid phosphatase by shaking and protection by added surface-active agent. Shaking mixtures (20 ml) contained purified enzyme (056 /ug of protein/ml) in 0.05 M acetate buffer at pH 5.5. The solutions were shaken in 50 ml volumetric flasks using a mechanical shaker (Burrell, model CC). Temperatures were maintained by immersion of the flasks in an appropriately set water bath. After specified intervals of shaking, duplicate 0.1 ml ahquots were removed into tubes containing Triton X-100. All tubes were assayed simultaneously, following the shaking procedure, with 0.05 M phenyl phosphate as substrate. Curve 1 Enzyme + Triton X-100 at 0°C and 29°C. Curve 2 Enzyme alone at 0°C. Curve 3 Enzyme alone at 29°C. From Tsuboi and Hudson (88).

Inhibition of Prostatic Acid Phosphatase by Various Hydroxycarboxylic... 

Fia. 7. Inhibition of prostatic acid phosphatase by D-(+)-tartaric acid. The reaction mixtures all contained equivalent amounts of the enzyme preparation, the indicated concentration of substrate (pH 5.0), 0.05 iff acetate buffer (pH 5.0), and tartaric acid (pH 5.0) total volume, 4.5 ml. Each point represents average values of determinations made with 5 X 10-5 iff and 10 X 10"5 iff tartaric acid except in the case of /3-glycerophosphate for which 1 X 10 iff and 2 X 10 iff tartaric acid was used. From Kilsheimer and Axelrod (4 ). 

Prostatic acid phosphatase is partially and reversibly inactivated by calcium ion (45). Anions such as chloride, bromide, and thiocyanate inhibit prostatic acid phosphatase competitively with regard to substrate as well as noncompetitively. A kinetic analysis by London et al. (46) indicates that the noncompetitive inhibition was related to changes in charge on the protein molecule. A variety of nonspecific anions accelerate thermal denaturation of the enzyme. The enzyme is quite sensitive to a number of electrolyte changes, but it is not clear whether these factors are involved in biological control. 

Highly purified prostatic acid phosphatase labeled with 14C has been obtained by incubation of slices of hypertrophic human gland with labeled amino acids 53). 

Lavallee and Rosenkrantz (54) studied the purification of dog prostatic acid phosphatase from prostatic secretion obtained from pilocarpine-stimulated dogs with cystopreputiostomy prostatic fistulas. A 45-... 

Prostatic acid phosphatase is reversibly inactivated by p-mercuri-benzoate and by Cu2+ and Fe3+ (59). In contrast to red cell acid phosphatase, prostatic acid phosphatase is only partially inactivated even after prolonged periods of incubation at high concentrations of p-mercuri-benzoate. Addition of cysteine to the p-mercuribenzoate-treated enzyme produces complete reactivation. Binding of SH groups by p-mercuri-benzoate renders the enzyme more labile to thermal denaturation, but no difference is obtained with surface inactivation (23). Similar partial inactivation with Cu2+ is also subject to reactivation. 

Prostatic acid phosphatase is irreversibly inhibited by reaction with iodine monochloride at pH 8.1. Figures 9 and 10 (60) show the effect of concentration of IC1 and the time course of the reaction. Very rapid inactivation occurred at concentrations of 0.05 vaM IC1. Further increase in the concentration of the reagent produced further inactivation but... 

When prostatic acid phosphatase is purified so that it is completely free of diesterase activity, it can be used in a variety of structural... 

The incubation digest (7.0 ml) contained 1 ml of 0.022 M phenyl phosphate 2.5 ml of 0.1 M acetate buffer, pH 5.0 0.5 ml of test enzyme solution and 3.0 ml of solutions of acceptors giving a final concentration as shown in the third column. Incubation time, 30 min. Digests were inactivated by 3.0 ml of 10% trichloroacetic acid solution and were analyzed for phenol and inorganic phosphate. In the case of the standard acceptor, 1,4-butanediol, the expected transfer product, 1,4-butanediol phosphate, was isolated in a yield of 35% from a large-scale experiment. The hydrolysis of this phosphate ester by prostatic acid phosphatase liberated approximately equimolar amounts of 1,4-butanediol and inorganic phosphate. 

Table XI (73) shows the Stokes radii and frictional ratio obtained by the study of purified acid phosphatase. The preparations show molecular homogeneity during filtration on Sephadex G-100, in the analytical ultracentrifuge, and during immunolectrophoresis. These data obtained by chromatography on Sephadex G-200 indicate that human prostatic acid phosphatase has an effective Stokes radius of 47.1 A and a frictional ratio of 1.56, suggesting considerable molecular asymmetry.

Shaw (115) reported a 300-fold purification of enzyme from tobacco leaves. Activity of the enzyme was optimal at pH 5.5-5.7, and divalent cations were not required for activity. The enzyme possessed high activity toward ribonucleoside 2 - and 5 -monophosphates and glucose 1-phosphate. There was no activity toward RNA or phosphodiesters. Fluoride acts as a noncompetitive inhibitor for this enzyme. This behavior of fluoride is in contrast to the behavior with prostatic acid phosphatase where the inhibition is strictly competitive. 

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