Purification and properties of oestrogen-induced uterine peroxidase.
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ABSTRACT:
SUBMITTER: McNabb T
PROVIDER: S-EPMC1172357 | biostudies-other | 1975 Nov
REPOSITORIES: biostudies-other
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Project description:The distribution of oestrogen-induced peroxidase in the resuspended 8000g pellet of rat uterine homogenates was examined by centrifugation in a sucrose density gradient. Within 10h of treatment with oestradiol, peroxidase activity was found in a region devoid of catalase or urate oxidase (peroxisomal markers) which did not overlap the fractions containing succinate dehydrogenase (mitochondrial marker) or acid phosphatase (lysosomal marker). The induced uterine enzyme was localized in reticular membrane-bound vesicles with isopycnic density of 1.28g/ml from which it could be released by treatment with detergent.
| S-EPMC1164228 | biostudies-other
Project description:1. An enzyme that catalyses the metabolism and binding of [4-(14)C]oestradiol to protein and to other high-molecular-weight substances in the presence of H(2)O(2) was shown to be absent from the uteri of immature rats and to be induced by physiological doses of oestrogen or pregnant-mare-serum gonadotrophin. 2. The pH optimum, stability to heat and other characteristics of the uterine enzyme system as well as its subcellular distribution were determined. 3. The increase in the ability of uterine preparations to convert [4-(14)C]oestradiol into water-soluble products as a result of oestrogen treatment was accompanied by an increase in peroxidase and NADH oxidase activities and was inhibited by actinomycin D and cycloheximide. 4. The results support the proposal that the increase in peroxidase activity after oestrogen treatment might be part of an adaptive response of the uterus permitting it to bind and inactivate oestrogens and thus limit the duration of their effect upon this target tissue.
| S-EPMC1178688 | biostudies-other
Project description:An oestrogen-induced secretory protein from mouse uterine luminal fluid was purified by CM-Affi-Gel Blue chromatography and reverse-phase h.p.l.c. This protein has an apparent molecular mass of approx. 70 kDa both by SDS/polyacrylamide-gel electrophoresis (with or without 2-mercaptoethanol) and by gel-filtration column chromatography, indicating that it exists as a single-chain polypeptide. Further analysis of the protein revealed that it is highly basic (pI greater than or equal to 10) and is a glycoprotein. The N-terminus appears to be blocked to Edman degradation. The partial amino acid sequence of a fragment was obtained by cleavage with CNBr; no sequence homology was apparent between the analysed fragment and other known sequences. The incorporation of [35S]methionine into uterine proteins in vitro revealed that oestrogen treatment of immature mice stimulates both synthesis and secretion of the 70 kDa protein. An enzyme-linked immunosorbent assay with polyclonal antibody was used to determine the tissue distribution of the protein. Tissues such as lung, brain, spleen, muscle, intestine, liver, kidney and ovary of oestrogen-treated mice did not have detectable amounts of the 70 kDa protein. Immunoreactivity was present in uterine and vaginal tissues from oestrogen-treated animals. The 70 kDa protein was not induced by testosterone or progesterone. Although the function of this protein is unknown, it is useful as a marker for the study of oestrogen action in the mammalian uterus as well as regulation of gene expression at the molecular level.
| S-EPMC1147433 | biostudies-other
Project description:Rat eosinophil peroxidase and rat uterine peroxidase II showed similar electrophoretic mobilities, molecular weights, specific activities and spectral properties and could be purified by essentially identical techniques. Antibodies raised against the uterine enzyme strongly inhibited the eosinophil enzyme. It is suggested that rat eosinophil peroxidase and rat uterine peroxidase II may well be one and the same enzyme.
| S-EPMC1153907 | biostudies-other
Project description:1. The visible absorption spectrum of peroxidase II, isolated from the uterine tissue of oestradiol-treated rats, and some of its derivatives were recorded. The spectral properties of this enzyme are very similar to eosinophile peroxidase and lactoperoxidase, suggesting that these enzymes may have a similar form of haem as prosthetic group. 2. The uterine peroxidase is modified upon interaction with H2O2 and the difference spectrum of this modified enzyme is similar to that of complex II of lactoperoxidase. The modified enzyme was found to revert spontaneously to the native enzyme at rates which depended on the concentration of free enzyme and H2O2.
| S-EPMC1163612 | biostudies-other
Project description:In this study, peroxidase from Ziziphus jujuba was purified using ion exchange, and gel filtration chromatography resulting in an 18.9-fold enhancement of activity with a recovery of 20%. The molecular weight of Z. jujuba peroxidase was 56?kDa, as estimated by Sephacryl S-200. The purity was evaluated by SDS, which showed a single prominent band. The optimal activity of the peroxidase was achieved at pH 7.5 and 50?°C. Z. jujuba peroxidase showed catalytic efficiency (Kcat/Km) values of 25 and 43 for guaiacol and H2O2, respectively. It was completely inactivated when incubated with ?-mercaptoethanol for 15?min. Hg2+, Zn2+, Cd2+, and NaN3 (5?mM) were effective peroxidase inhibitors, whereas Cu2+ and Ca2+ enhanced the peroxidase activity. The activation energy (Ea) for substrate hydrolysis was 43.89?kJ?mol-1, while the Z value and temperature quotient (Q10) were found to be 17.3?°C and 2, respectively. The half-life of the peroxidase was between 117.46 and 14.15?min. For denaturation of the peroxidase, the activation energy for irreversible inactivation Ea*(d) was 120.9 kJmol-1. Thermodynamic experiments suggested a non-spontaneous (?G*d > 0) and endothermic reaction phase. Other thermodynamic parameters of the irreversible inactivation of the purified enzyme, such as ?H* and ?S*, were also studied. Based on these results, the purified peroxidase has a potential role in some industrial applications.
| S-EPMC7224213 | biostudies-literature
Project description:Glutathione peroxidase (glutathione--H2O2 oxidoreductase; EC 1.11.1.9) was purified to homogeneity from human placenta by using (NH4)2SO4 precipitation, ion-exchange chromatography, Sephadex gel filtration and preparative polyacrylamide-disc-gel electrophoresis. Glutathione peroxidase from human placenta is a tetramer, having 4g-atoms of selenium/mol of protein. The molecular weight of the enzyme is about 85000 with a subunit size of about 22,000. Kinetic properties of the enzyme are described. On incubation with cyanide, glutathione peroxidase is completely and irreversibly inactivated and selenium is released as a low-molecular-weight fragment. Reduced glutathione, beta-mercaptoethanol and dithiothreitol protect the enzyme from inactivation by cyanide and the release of selenium. Properties of human placental glutathione peroxidase are similar to those of isoenzyme A reported earlier by us from human erythrocytes. The presence of isoenzyme, B, reported earlier by us in human erythrocytes, was not detected in placenta. Also selenium-independent glutathione peroxidase (isoenzyme II), which is specific for cumene hydroperoxide, was not present in human placenta.
| S-EPMC1186396 | biostudies-other
Project description:IntroductionUterine fibroids (UFs) are benign, monoclonal tumours of the female genital tract that originate from the myometrium. They may be diagnosed in as many as 80% of women depending on the selected population. UFs depend mostly on steroid hormones. Elevated levels of oestrogens and progesterone are believed to be among the most important factors inducing their formation and growth. These facts suggest that oestrogen (ESR) and progesterone receptors are crucial in UF pathophysiology as well. Previous studies have shown that, in some populations, polymorphisms in ESR genes (e.g. PvuII) constitute an important risk factor for UFs.Material and methodsThe aim of our study was to investigate whether ESRα PvuII polymorphism is associated with an increased risk of UFs in Caucasian women of Polish origin. A total of 197 patients (114 UF-positive and 83 controls) were included in this retrospective cohort study. ESRα gene polymorphism PvuII (rs2234693) was assayed with PCR and restriction fragment length polymorphism (RFLP).ResultsOur study found no significant difference in the occurrence of ESR PvuII polymorphism between women with UFs and UF-free controls in the selected population.ConclusionsOur results did not indicate a significant association between ESRα gene PvuII polymorphism and the risk of UFs in Caucasian women of Polish origin. More studies and comparisons between races are necessary to clarify the role of ESRα in the development and progression of UFs.
| S-EPMC6372853 | biostudies-literature
Project description:Myeloperoxidase and eosinophil peroxidase have been isolated from outdated human blood. Peroxidase activity was extracted from washed leucocytes using 0.5 M-CaCl2 and the extract further purified by chromatography on concanavalin A--Sepharose, phenyl-Sepharose and finally by gel filtration. The final enzyme preparations were highly purified according to spectral and gel-electrophoretic criteria. Under reducing and denaturing conditions on polyacrylamide-gel electrophoresis myeloperoxidase gave rise to bands of Mr 57 000, 39 000 and 15 500, whereas the eosinophil enzyme yielded bands of Mr 50 000 and 15 500. Both enzymes were very resistant to denaturation either by the chaotropic agents urea and guanidinium chloride or by elevated temperatures. Spectral properties of the native and reduced forms of the enzymes are reported.
| S-EPMC1154157 | biostudies-other
Project description:Human papillomavirus (HPV) is required but not sufficient for cervical carcinoma (CxCa) development. Oestradiol (E2 ) promotes CxCa development in K14E7 transgenic mice expressing the HPV16 E7 oncoprotein under the control of the keratin (K14) promoter. E2 mainly functions through oestrogen receptor ? (ER?). However, the role of ER? in human CxCa has been underappreciated largely because it is not expressed in carcinoma cells. We have shown that deletion of Esr1 (the ER?-coding gene) in the cervical stroma of K14E7 mice promotes regression of cervical intraepithelial neoplasia (CIN), the precursor lesion of CxCa. Here, we deleted Esr1 in the cervical epithelium but not in the stroma. We found that E2 induced cervical epithelial cell proliferation in epithelial ER?-deficient mice. We also found that E2 promoted the development of CIN and CxCa in epithelial ER?-deficient K14E7 mice and that all neoplastic epithelial cells were negative for ER?. In addition, proliferation indices were similar between ER?- and ER?+ CxCa. These results indicate that epithelial ER? is not necessary for E2 -induced CIN and CxCa. Taking these findings together, we conclude that stromal ER? rather than epithelial ER? mediates oncogenic E2 signalling in CxCa. Our results support stromal ER? signalling as a therapeutic target for the disease. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
| S-EPMC5959269 | biostudies-literature