Tartrate prostatic

Red blood cells also contain sufficient acid phenylphospha-tase for mild hemolysis to cause false elevations. Therefore, inhibitors such as ethanol, formaldehyde, copper sulfate> and 1-tartrate have been used to inhibit selectively the enzyme of one or more tissues and enhance the specificity of the test (101). Ethanol is unsuitable because it inhibits the enzyme from erythrocytes and prostate simultaneously, and because it yields serum activities which correlate poorly with prostatic disease. Formaldehyde inhibits the erythrocytic enzyme and has been said to yield clinically satisfactory results. The copoper resistant acid phosphatase of serum is elevated by metastatic carcinoma of the breast, as well as by other metastatic cancers, and is also elevated by a wide variety of non-cancerous diseases. 

Tartrate is a most widely used inhibitor of prostatic acid phenylphosphatase activity. With this inhibitor Bonner and associates detected five cases of unsuspected carcinoma of the prostate in 221 hospital patients and clarified the diagnoses in another four patients (98). Nonetheless, the diagnostic specificity of an elevated 1-tartrate inhibited activity is not absolute, as Fishman et. al found it elevated in 48 of 1,190 males without cancer. Whitmore and associates also observed this activity to be elevated in 3 of 20 patients with uncomplicated nodular hyperplasia (102). Hill compared the total versus the "prostatic" serum activities in 20 patients with localized untreated carcinoma of the prostate and observed... 

Acid phosphatases are produced by erythrocytes, the liver, kidney, spleen, and prostate gland. The enzyme of the prostate gland is clinically important, because its increased activity in the blood can be an indication of prostate cancer. The phosphatase from the prostate gland is strongly inhibited by tartrate ion, but acid phosphatases from other tissues are not. How can this information be used to develop a specific procedure for measuring the activity of the acid phosphatase of the prostate gland in human blood serum ... 

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. 

Answer First, measure the total acid phosphatase activity in a blood sample in units of /nnol of phosphate ester hydrolyzed per mL of serum. Next, remeasure this activity in the presence of tartrate ion at a concentration sufficient to completely inhibit the enzyme from the prostate gland. The difference between the two activities represents the activity of acid phosphatase from the prostate gland. 

Abul-Fadl and King (Al, A2, A3, A4) also investigated the effect of various ions and organic compounds on the acid phosphatase activity of these two tissues. Without describing the results in detail, some of the outstanding effects may be noted. A concentration of 0.5 X 10 M Cu inhibited erythrocytic phosphatase to the extent of 88-96%, but prostatic phosphatase only to the extent of 10-18%. Similarly, 0.5% formaldehyde inhibited completely the erythrocytic phosphatase, but had no effect on prostatic phosphatase. The reverse patterns were shown by 0.5 X 10 M Fe + (ferric) ion, which inhibited erythrocytic phosphatase slightly, about 5-9%, and inhibited the prostatic enzyme to the extent of 80%. Fluoride in 0.01 Af concentration also had comparatively little effect (8% inhibition) on erythrocytic phosphatase but exerted a marked inhibition, 96%, on prostatic phosphate. Of various organic radicals tested, only L-( + )-tartrate (0.01 A/) had a marked differential effect, with 94% inhibition of the prostatic phosphatase and none of the erythrocytic phosphatase. 

There is no information concerning the isoenzymatic composition of the purified prostatic phosphatases that were used in the preceding kinetic studies. Nor do there appear to be any kinetic studies on the individual isoenzymes. The possibility exists that substantial differences in kinetic characteristics, such as the value for Ki, for l-(4-)-tartrate. 

Another procedure to increase the specificity of acid phosphatase determinations for prostatic disease has involved the use of n- (-I-) -tartrate to distinguish between the enzyme from the prostate and other tissues. In a series of papers from 1947 to 1949, Abul-Fadl and King (Al, A2, A3, A4) studied the properties of various acid phosphatases and reported that 0.01 Af L- (4-) -tartrate inhibited the hydrolysis of phenyl phosphate by human prostatic acid phosphatase dissolved in normal saline or in plasma to the extent of 95%, but had no effect on the hydrolysis by acid phosphatase from erythrocytes. The inhibitions of acid phosphatases from other human tissues were as follows liver, 70% kidney, 80% spleen, 70%. 

Several years later, Fishman and his associates (FI, F2, F3) applied this principle to the determination of the tartrate-inhibitable or prostatic fraction in serum. This method involved the hydrolysis of disodium phenyl phosphate into phenol and phosphate by serum in the absence and presence of 0.02 Af n- (4-) -tartrate for a period of 1 hour at 37°C. Suitable blank and control solutions were employed. The activity in the... 

Acetoacetic acid,p-aminosalicylic acid, gentisic acid, pansporin, and bilirubin react with diazo dye Fast Red TR Salt. The addition of sodium tartrate to a final concentration of 0.05 mol/1 (pH S.S.) inhibits the prostatic isoenzyme of acid phosphatase and permits the measurement of the result due primarily to these interferents. AcP in samples suspected to contain these interferents may be analyzed before and after treatment with sodium tartrate to get the AcP and Blank results, respectively. 

Although once widely used to detect or monitor carcinoma of the prostate, determination of ACP (tartrate-inhibited) activity in serum is now rarely used for this purpose. It has been replaced by prostate-specific antigen (PSA). 

Continuous-monitoring methods for assay of TR-ACP activity are based on the principle introduced by Hillmann in which a-naphthoi released from its phosphate ester forms a colored product with the stabilized diazonium salt of 2-amino-5-chlorotoluene-1,5-naphthalene disulfonate (Fast Red TR). The introduction of alcohols, such as 1,5-pen-tanediol, accelerates the reaction and increases sensitivity by acting as phosphate acceptors in transfer reactions. The addition of sodium tartrate inhibits the sensitive isoenzymes (i.e., prostatic and lysosomal ACPs) if they are present in the sample. 

Acid phosphatases are enzymes that have been studied extensively due to the fact that their dysregulation is associated with pathophysiological conditions. This characteristic has been exploited for the development of diagnostic and therapeutic methods. As an example, prostatic acid phosphatase was the first marker for metastatic prostate cancer diagnosis and the dysregulation of tartrate resistant acid phosphatase is associated with abnormal bone resorption linked to osteoporosis. 

The pioneering crystallization smdies on prostatic acid phosphatase and mammalian tartrate-resistant acid phosphatase conformed significant milestones towards the elucidation of the mechanisms followed by these enzymes (Schneider et al., EMBO J 12 2609-2615, 1993). Acid phosphatases are also found in nonmammalian species such as bacteria, fungi, parasites, and plants, and most of them share structural similarities with mammalian acid phosphatase enzymes. 

Key words Prostate cancer, Bone resorption, Osteoporosis, MetaUo-enzymes, Tartrate-resistant... 

The lysosomal acid phosphatase enzyme played a key role in the discovery of lysosomes by de Duve in 1963 and is widely used as a lysosomal marker. This enzyme shows a high degree of sequence similarity (ca. 49 % identity) with prostatic acid phosphatase [9] and both are inhibited by L-(+)-tartrate ion [10]. 

Prostatic acid phosphatase Phosphate ( P04 ), vanadate ( ), arsenate ( AsO ), molybdate ( MoO ), L-(+(-tartrate, benzylaminophosphonic acid and benzylaminobenzylphosphonic acid, W-propyltartramate Aliphatic alcohols... 

High-scale purification methods are required for several protein studies such as crystallography, mass specttometty, circular dichroism, and function. Here we describe a purification method for PAP based on anion exchange, L-(+)-tartrate affinity, and gel filtration chromatographies. Acid phosphatase activity and protein concentration were measured for each purification step, and to collect the firactions with the highest acid phosphatase activity the p-nittophenyl phosphate method was used. The purified protein obtained by the procediue described here was used for the determination of the first reported three-dimensional structure of prostatic add phosphatase. 

Key words Prostatic acid phosphatase. Recombinant protein. High-scale purification, Afimity chromatography, L-(-r)-tartrate, Acid phosphatase activity, p-Nitrophenyl phosphate... 

Prostatic acid phosphatase (EC 3.1.3.2) can be purified from seminal fluid, prostate tissue, or as a recombinant protein. High-scale puriflcation methods are important in order to obtain mass amounts of homogeneous, purifled protein required by structural and functional studies such as inhibitor and activator analyses. Acid phosphatases can be divided into two groups according to their sensitivity or resistance to L-(-i-)-tartrate inhibition. The activity of these enzymes can be analyzed by the -nitrophenyl phosphate method in presence [1] or absence of L-(-i-)-tartrate [2]. PAP belongs to the group of L-(-i-)-tartrate sensitive acid phosphatases, and this feature is used as the basis of the puriflcation method described in this chapter. 

Small-scale protein purification protocol is suitable when seminal fluid or prostate tissue is used as starting material. Perform all the steps for small-scale protein purification in +4 °C (or cold room). Same buffers than in mass-scale purification are used. Prepare your own columns using strong anion exchanger matrix such as QAE Sephadex A-25, and for gel filtration matrix use e.g. Sephacryl S-200 HR. L-(+)-tartrate afiSnity column is prepared in the same way than for mass-scale purification method. Use peristaltic pumps, gradient mixer and fraction collector in +4 °C. 

Nncleotidase Acid lahUe snhunit of IGFBP Acid phophatase, tartrate resistant Acid phosphatase, prostatic Actin heta (from platelets)... 

Determination of the acid tartrate-sensitive P. in serum is important in the diagnosis of prostate cancer, which is accompanied by a marked increase in the serum level of this enzyme. Measurement of alkaline P in serum is used in the diagnosis of bone disease, especially bone tumors, and diseases of the liver (hepatitis) and the gall bladder (e.g. obstructive jaundice), all of which result in a several fold increase in serum P. The isoenzymes of both acid and alkaline P. differ in their neuraminic acid contents. The most widely used substrate for P. assay is p-nitrophenyl phosphate the released p-nitrophenol can be determined directly by photometry (k , 405 nm). 

A comparison with the acid phosphatase (s) which may be increased in carcinoma of the prostate, Paget s disease, certain liver diseases, and hyperparathyroidism showed that the acid phosphatase of Gaucher s disease can be differentiated from the former with the use of various activators and inhibitors (Grundig et al. 1965). In contrast to prostatic phosphatases it is not inhibited by L-tartrate... 

Most of the substrates that have been used in measuring alkaline phosphatase activity have also been used to measure acid phosphatase, e.g. p-nitrophenylphosphate, phenylphosphate and a-naphthyl phosphate. In most cases of acid phosphatase estimation, it is the level of the prostatic phosphatase which needs to be known, and so specific inhibitors are included in the reaction mixtures. The prostatic isoenzyme is strongly inhibited by tartrate and so in many methods it is tartrate-labile acid phosphatase which is measured. On the pther hand, the red cell enzyme which contributes significantly to the total serum activity is inhibited by formaldehyde and cupric ions. Many laboratories therefore measure formaldehyde-stable acid phosphatase as an indication of prostatic acid phosphatase. 

---