scholarly journals MICROSOMAL AND LYSOSOMAL ACID PHOSPHATASE ISOENZYMES OF MOUSE KIDNEY CHARACTERIZATION AND SEPARATION

1972 ◽  
Vol 20 (7) ◽  
pp. 487-498 ◽  
Author(s):  
CHI-WEI LIN ◽  
WILLIAM H. FISHMAN
Keyword(s):  
Acid Phosphatase ◽  
Migration Rate ◽  
Biochemical Properties ◽  
Mouse Kidney ◽  
Acid Phosphatases ◽  
Ph Optimum ◽  
Lysosomal Acid ◽  
Lysosomal Fraction ◽  
Heat Stable ◽  
Diethylaminoethyl Cellulose

Biochemical methods have been employed to characterize and separate acid phosphatases from lysosomal and microsomal fractions in order to decide whether different isoenzymes reside in these subcellular locations. Microsomal and lysosomal fractions of mouse kidney homogenate were isolated by differential centrifugation. Acid phosphatase of lysosomal fraction goes into solution after lysosomes have been repeatedly frozen and thawed, whereas acid phosphatase of microsomal fraction is firmly bound to the membrane and is freed of contamination by lysosomal enzyme after ultrasonication and centrifugation. The membrane-bound microsomal acid phosphatase is labile at 37°C, pH 4.9, more active toward phenolic substrates (phenyl phosphate and p-nitrophenyl phosphate) than β-glycerophosphate, α-naphthol phosphate or naphthol AS-BI phosphate. It also has a higher pH optimum (6.3), is resistant to l-tartrate and oxalate inhibition and has a slower electrophoretic migration rate in Triton X-100-impregnated polyacrylamide gels. The free lysosomal acid phosphatase is relatively heat-stable, is less active against phenolic substrates, is sensitive to l-tartrate and oxalate inhibition, has a lower pH optimum (5.6) and has a faster migration rate in electrophoresis. These two acid phosphatases can also be separated by diethylaminoethyl-cellulose chromatography. This study thus demonstrated the existence of an acid phosphatase isoenzyme in the microsomal membrane with different biochemical properties from the lysosomal isoenzyme of acid phosphatase.

The Demonstration of Human Prostatic Acid Phosphatase (Pap) With Phosphorylcholine

1976 ◽  
Vol 34 ◽  
pp. 88-89
Author(s):  
José A. Serrano ◽  
Hannah L. Wasserkrug ◽  
Anna A. Serrano ◽  
Arnold M. Seligman
Keyword(s):  
Acid Phosphatase ◽  
Prostate Gland ◽  
Prostatic Acid Phosphatase ◽  
Human Prostate ◽  
Rat Tissues ◽  
Specific Substrate ◽  
Acid Phosphatases ◽  
Lysosomal Acid ◽  
Cacodylate Buffer

As previously reported (1, 2) phosphorylcholine (PC) is a specific substrate for prostatatic acid phosphatase (PAP) as opposed to other acid phosphatases, e.g., lysosomal acid phosphatase. The specificity of PC for PAP is due to the pentavalent nitrogen in PC, a feature that renders PC resistant to hydrolysis by all other acid phosphatases. Detailed comparative cytochemical results in rat tissues are in press. This report deals with ultracytochemical results applying the method to normal and pathological human prostate gland.Fresh human prostate was obtained from 7 patients having transurethral resections or radical prostatectomies. The tissue was fixed in 3% glutaraldehyde- 0.1 M cacodylate buffer (pH 7.4) for 15 min, sectioned at 50 μm on a Sorvall TC-2 tissue sectioner, refixed for a total of 2 hr, and rinsed overnight in 0.1 M cacodylate buffer (pH 7.4)-7.5% sucrose.

scholarly journals ISOZYMES OF ACID PHOSPHATASE IN NORMAL AND CALMETTE-GUÉRIN BACILLUS-INDUCED RABBIT ALVEOLAR MACROPHAGES

1968 ◽  
Vol 128 (5) ◽  
pp. 1031-1048 ◽  
Author(s):  
S. G. Axline
Keyword(s):  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Alveolar Macrophages ◽  
Acid Phosphatase Activity ◽  
Enzyme Inhibitors ◽  
Freezing And Thawing ◽  
Ph Optimum ◽  
Lysosomal Fraction ◽  
Repeated Freezing ◽  
Major Acid

The acid phosphatase activity of normal alveolar and BCG-induced alveolar macrophages has been examined. Five electrophoretically distinct forms of acid phosphatase have been identified in both normal and BCG-induced macrophages. The acid phosphatases can be divided into two major categories. One category, containing four distinct forms, is readily solubilized after repeated freezing and thawing or mechanical disruption The second category, containing one form, is firmly bound to the lysosomal membrane and can be solubilized by treatment of the lysosomal fraction with Triton X-100. The Triton-extractable acid phosphatase and the predominant aqueous soluble acid phosphatase have been shown to differ in the degree of membrane binding, in solubility, in net charge, and in molecular weight. The two pre-dominant phosphatases possess identical pH optimum and do not differ in response to enzyme inhibitors. BCG stimulation has been shown to result in a nearly twofold increase in acid phosphatase activity. A nearly proportionate increase in the major acid phosphatase forms has been observed.

LINGCOD MUSCLE PHOSPHOMONOESTERASES: I. ACID PHOSPHATASES THAT HYDROLYZEp-NITROPHENYL PHOSPHATE

1960 ◽  
Vol 38 (1) ◽  
pp. 605-612 ◽  
Author(s):  
Neil Tomlinson ◽  
R. A. J. Warren
Keyword(s):  
Metal Ions ◽  
The Other ◽  
Acid Phosphatases ◽  
Free Base ◽  
Ph Optimum ◽  
Ophiodon Elongatus ◽  
Nitrophenyl Phosphate ◽  
Diethylaminoethyl Cellulose ◽  
Ph Range ◽  
Acid Phosphomonoesterase

Five fractions (A to E), each possessing acid phosphomonoesterase activity, were separated from an aqueous extract of the muscle of lingcod (Ophiodon elongatus) by stepwise chromatography on diethylaminoethyl cellulose in the free-base form.Fraction A required Zn++or Mn++for activity, was inhibited by heparin, and had its pH optimum at 6.0. Fraction E required Zn++for activity, was not inhibited by heparin, and had its pH optimum at 5.5. Fractions B, C, and D did not require metal ions for activity, and were distinguished from each other by differences in response to pH, cysteine, ethylenediaminetetraacetate, fluoride, and tartrate.The pH range over which fraction A was active was shifted to slightly higher values when Mn++was the activator rather than Zn++. Also, A was inhibited strongly by cysteine when activated by Zn++, but not when activated by Mn++. Data are presented that indicate these differences were due to different properties of the activating ions, rather than to the presence in fraction A of two enzymes, one activated by Zn++and the other by Mn++.

Overlapping functions of lysosomal acid phosphatase (LAP) and tartrate-resistant acid phosphatase (Acp5) revealed by doubly deficient mice

Development ◽  
2001 ◽  
Vol 128 (23) ◽  
pp. 4899-4910
Author(s):  
Anke Suter ◽  
Vincent Everts ◽  
Alan Boyde ◽  
Sheila J. Jones ◽  
Renate Lüllmann-Rauch ◽  
...  
Keyword(s):  
Acid Phosphatase ◽  
Tartrate Resistant Acid Phosphatase ◽  
Arylsulfatase A ◽  
Acid Phosphatases ◽  
Lysosomal Storage ◽  
Lysosomal Acid ◽  
Biochemical Analyses ◽  
Filamentous Material ◽  
Gait Disturbances ◽  
Deficient Mice

To date, two lysosomal acid phosphatases are known to be expressed in cells of the monocyte/phagocyte lineage: the ubiquitously expressed lysosomal acid phosphatase (LAP) and the tartrate-resistant acid phosphatase-type 5 (Acp5). Deficiency of either acid phosphatase results in relatively mild phenotypes, suggesting that these enzymes may be capable of mutual complementation. This prompted us to generate LAP/Acp5 doubly deficient mice. LAP/Acp5 doubly deficient mice are viable and fertile but display marked alterations in soft and mineralised tissues. They are characterised by a progressive hepatosplenomegaly, gait disturbances and exaggerated foreshortening of long bones. Histologically, these animals are distinguished by an excessive lysosomal storage in macrophages of the liver, spleen, bone marrow, kidney and by altered growth plates. Microscopic analyses showed an accumulation of osteopontin adjacent to actively resorbing osteoclasts of Acp5- and LAP/Acp5-deficient mice. In osteoclasts of phosphatase-deficient mice, vacuoles were frequently found which contained fine filamentous material. The vacuoles in Acp5- and LAP/Acp5 doubly-deficient osteoclasts also contained crystallite-like features, as well as osteopontin, suggesting that Acp5 is important for processing of this protein. This is further supported by biochemical analyses that demonstrate strongly reduced dephosphorylation of osteopontin incubated with LAP/Acp5-deficient bone extracts. Fibroblasts derived from LAP/Acp5 deficient embryos were still able to dephosphorylate mannose 6-phosphate residues of endocytosed arylsulfatase A. We conclude that for several substrates LAP and Acp5 can substitute for each other and that these acid phosphatases are essential for processing of non-collagenous proteins, including osteopontin, by osteoclasts.

Immunological and biochemical evidence for identity of tartrate-resistant isoenzymes of acid phosphatases from human serum and tissues.

1980 ◽  
Vol 26 (3) ◽  
pp. 420-422 ◽  
Author(s):  
K W Lam ◽  
P Lee ◽  
C Y Li ◽  
L T Yam
Keyword(s):  
Acid Phosphatase ◽  
Substrate Specificity ◽  
Human Serum ◽  
Giant Cell ◽  
Bone Tumor ◽  
Biochemical Characterization ◽  
Acid Phosphatases ◽  
Ph Optimum ◽  
Human Spleen ◽  
Biochemical Evidence

Abstract We purified acid phosphatase isoenzyme 5b from a human spleen affected by leukemic reticuloendotheliosis and used it to produce a specific antiserum. The antiserum was used to show complete immunological identity among isoenzymes 5a and 5b in human serum, and 5b isolated from a giant-cell bone tumor and from the spleen of a case of Hodgkin's disease. Acid phosphatase 5b in a giant-cell bone tumor was isolated for biochemical characterization. Its pH optimum and substrate specificity were very similar to those of isoenzyme 5b from human spleen.

Immunological and biochemical evidence for identity of tartrate-resistant isoenzymes of acid phosphatases from human serum and tissues.

1980 ◽  
Vol 26 (3) ◽  
pp. 420-422 ◽  
Author(s):  
K W Lam ◽  
P Lee ◽  
C Y Li ◽  
L T Yam
Keyword(s):  
Acid Phosphatase ◽  
Substrate Specificity ◽  
Human Serum ◽  
Giant Cell ◽  
Bone Tumor ◽  
Biochemical Characterization ◽  
Acid Phosphatases ◽  
Ph Optimum ◽  
Human Spleen ◽  
Biochemical Evidence

Abstract We purified acid phosphatase isoenzyme 5b from a human spleen affected by leukemic reticuloendotheliosis and used it to produce a specific antiserum. The antiserum was used to show complete immunological identity among isoenzymes 5a and 5b in human serum, and 5b isolated from a giant-cell bone tumor and from the spleen of a case of Hodgkin's disease. Acid phosphatase 5b in a giant-cell bone tumor was isolated for biochemical characterization. Its pH optimum and substrate specificity were very similar to those of isoenzyme 5b from human spleen.

scholarly journals LYSOSOMAL ACID HYDROLASES IN MICE INFECTED WITH BCG

1965 ◽  
Vol 121 (5) ◽  
pp. 727-738 ◽  
Author(s):  
Kazuhisa Saito ◽  
Emanuel Suter
Keyword(s):  
Acid Phosphatase ◽  
Liver Homogenate ◽  
Subcellular Fractions ◽  
The Other ◽  
Acid Phosphatases ◽  
Acid Hydrolases ◽  
Lysosomal Acid

Experiments are reported dealing with the increase of lysosomal acid hydrolases induced by BCG infection. Acid hydrolases were determined quantitatively in peritoneal MP, liver homogenate, and plasma of normal and BCG-infected mice. A significant increase of acid phosphatase, ß-glucuronidase, and cathepsin was found in MP and liver homogenate of BCG-infected mice. In plasma also a significant increase of acid phosphatase and ß-glucuronidase was noticed. The results of the determination of the enzymes in centrifugally separated subcellular fractions of liver homogenate indicated clearly that the acid hydrolases associated mainly with the "large granular" fraction, which consists of mitochondria, lysosomes, and microsomes and that infection with BCG caused significant increase of the enzymes specifically in this fraction. Differences in the pattern of location among centrifugally separated fraction of liver homogenate were observed between acid phosphatase and the other two acid hydrolases. MP cultured in vitro doubled their acid phosphatases content within 24 hours, whereas ß-glucuronidase rather decreased in the same cells.

LINGCOD MUSCLE PHOSPHOMONOESTERASES: I. ACID PHOSPHATASES THAT HYDROLYZEp-NITROPHENYL PHOSPHATE

1960 ◽  
Vol 38 (6) ◽  
pp. 605-612 ◽  
Author(s):  
Neil Tomlinson ◽  
R. A. J. Warren
Keyword(s):  
Metal Ions ◽  
The Other ◽  
Acid Phosphatases ◽  
Free Base ◽  
Ph Optimum ◽  
Ophiodon Elongatus ◽  
Nitrophenyl Phosphate ◽  
Diethylaminoethyl Cellulose ◽  
Ph Range ◽  
Acid Phosphomonoesterase

Five fractions (A to E), each possessing acid phosphomonoesterase activity, were separated from an aqueous extract of the muscle of lingcod (Ophiodon elongatus) by stepwise chromatography on diethylaminoethyl cellulose in the free-base form.Fraction A required Zn++or Mn++for activity, was inhibited by heparin, and had its pH optimum at 6.0. Fraction E required Zn++for activity, was not inhibited by heparin, and had its pH optimum at 5.5. Fractions B, C, and D did not require metal ions for activity, and were distinguished from each other by differences in response to pH, cysteine, ethylenediaminetetraacetate, fluoride, and tartrate.The pH range over which fraction A was active was shifted to slightly higher values when Mn++was the activator rather than Zn++. Also, A was inhibited strongly by cysteine when activated by Zn++, but not when activated by Mn++. Data are presented that indicate these differences were due to different properties of the activating ions, rather than to the presence in fraction A of two enzymes, one activated by Zn++and the other by Mn++.

scholarly journals Heterogeneity of liver acid phosphatases in developing chick embryo

1969 ◽  
Vol 115 (2) ◽  
pp. 191-197 ◽  
Author(s):  
K.-M. Wang
Keyword(s):  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Acid Phosphatase Activity ◽  
Soluble Fraction ◽  
The Other ◽  
Acid Phosphatases ◽  
Peak Activity ◽  
Ph Optimum ◽  
Triton X ◽  
Soluble Fractions

1. The development, localization and heterogeneity of acid phosphatase and a Zn2+-activated acid phosphatase in cellular fractions of developing chick liver were studied. 2. Acid phosphatase is distributed abundantly in the particulate and soluble fractions. The soluble fraction is rich in Zn2+-activated acid phosphatase, which attains its peak activity at about 15 days of incubation. 3. The particulate acid phosphatase activity is inhibited by fluoride but not by sodium l(+)-tartrate or cysteine. On the other hand, the soluble Zn2+-activated acid phosphatase activity is inhibited by sodium l(+)-tartrate and cysteine but not by fluoride. 4. The pH optimum of these two enzymes is similar at about 5·6. 5. The soluble Zn2+-activated acid phosphatase activity appears to be thermally stabilized by the treatment with Triton X-100 or bovine serum albumin.

The acid phosphatases of bovine leucocytes, plasma and the milk of healthy and mastitic cows

1975 ◽  
Vol 42 (3) ◽  
pp. 391-400 ◽  
Author(s):  
A. T. Andrews ◽  
E. Alichanidis
Keyword(s):  
Molecular Weight ◽  
Acid Phosphatase ◽  
Exchange Resin ◽  
Bovine Milk ◽  
Ion Exchange Resin ◽  
Acid Phosphatases ◽  
Ph Optimum ◽  
Nitrophenyl Phosphate ◽  
Competitive Inhibitors ◽  
Electrophoretic Component

SummarySome of the acid phosphatase isozymes of bovine leucocytes and plasma have been separated and partly characterized. About 80% of the phosphatase activity of leucocytes at pH 4·9 was particle-bound and about 8% was extractable with Amberlite CG-50 ion exchange resin. This extractable enzyme existed as a single electrophoretic component with a mol. wt of about 42000 and with optimum activity at pH 5·8. Km for p-nitrophenyl phosphate was 1·6 mM at pH 5·8 and 0·4 mM at pH 4·9. At pH 5·8 orthophosphate (K1 = 1·5 mM) and pyrophosphate (Ki = 4·1 mM) were competitive inhibitors. The enzyme was also strongly inhibited by F−, Al3+, IO4− and S2032−. The enzyme which was not extractable with Amberlite was very heterogeneous with respect to molecular weight. At the pH optimum (4·9), Km for p-nitrophenyl phosphate was 0·4 mM and orthophosphate (K1 = 2·3 mM) and pyrophosphate (K1 = 2·1 mM) were competitive inhibitors. Other inhibitors included F−, Al3+, Hg2+, IO4− and tartrate. The enzyme extracted from plasma by Amberlite CG-50 treatment had properties similar to that extracted from leucocytes. Normal bovine milk contained a single acid phosphatase, but milk from cows with mastitis showed 3 electrophoretic isozyme bands, one being the same as in normal milk; the 2 additional bands were of leucocyte origin.

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