DNA-Polymerases α, δ and ε from T-Cell Spontaneous Lymphoblastic Leukemia of Sprague-Dawley Inbred Rat: Isolation and Characterization

1995 ◽  
Vol 60 (9) ◽  
pp. 1555-1572 ◽  
Author(s):  
Pavel Kramata ◽  
Jaroslav Černý ◽  
Gabriel Birkuš ◽  
Ivan Votruba ◽  
Berta Otová ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Lymphoblastic Leukemia ◽  
Dna Polymerases ◽  
Sedimentation Coefficient ◽  
Nuclear Antigen ◽  
Exonuclease Activity ◽  
Sprague Dawley ◽  
Dna Primase ◽  
Isolation And Characterization ◽  
Inbred Rat

Using a single isolation procedure and selective assays for the determination of enzyme activity, highly purified DNA-polymerases α, δ and ε were isolated from the lymphoma of Sprague-Dawley inbred rats. For pol α the subunit composition was 170, 70, 57 and 53 kDa with sedimentation coefficient 8.7 S for the native molecule; pol delta consists of two polypeptides (133 and 46 kDa; sedimentation coefficient 8.2 S), while pol ε is a single polypeptide (140 kDa) and its sedimentation coefficient is 7.0 S. Comparison of the interaction of individual enzymes with known inhibitors and proliferating cell nuclear antigen (PCNA) using the template-primer poly dA-oligo dT12-18, gave the following data: (i) pol α is selectively inhibited by N2-(p-butylphenyl)-2'-deoxyguanosine 5'-triphosphate (BuPdGTP) and stimulated by dimethyl sulfoxide; (ii) all the enzymes are inhibited by N-ethylmaleimide and aphidicolin; (iii) PCNA stimulates pol δ approximately 50 times while pol ε is moderately inhibited; (iv) pol α exhibits considerably higher DNA-primase activity with poly dC as template than with poly dT, and negligible 3'-5'-exonuclease activity whereas pol δ and pol ε, which do not exert any DNA-primase activity have approximately the same 3'-5'-exonuclease activity. The ability of individual polymerases to utilize poly dT-oligo dA12-18 as a template-primer at different pH values, ionic strengths and Mg2+-concentrations was also investigated. In comparison to poly dA-oligo dT12-18 template-primer, pol α has 140% of enzyme activity on poly dT-oligo dA12-18 under optimal conditions, whereas the activity of pol ε and pol δ is 4 times and 10 times lower, respectively.

Nonproteolyzed Form of DNA-Polymerase ε from T-Cell Spontaneous Lymphoma of Sprague-Dawley Inbred Rat: Isolation and Characterization

1998 ◽  
Vol 63 (5) ◽  
pp. 723-731 ◽  
Author(s):  
Gabriel Birkuš ◽  
Pavel Kramata ◽  
Ivan Votruba ◽  
Berta Otová ◽  
Miroslav Otmar ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Sedimentation Coefficient ◽  
Isolation Procedure ◽  
Sprague Dawley ◽  
Nucleotide Analogs ◽  
Isolation And Characterization ◽  
Inbred Rats ◽  
Catalyzed Reactions ◽  
Inbred Rat

Using a simple isolation procedure and selective assay for the determination of enzyme activity the nonproteolyzed and proteolyzed form of DNA-polymerase ε (pol ε and pol ε*) from the lymphoma of Sprague-Dawley inbred rats were purified. Nonproteolyzed pol ε is composed of two subunits (240 000 and 50 000) with sedimentation coefficient 10.5 S, while the subunit composition of pol ε* was 145 000 and 73 000. Estimated Km values for dATP and dGTP as well as Ki values for acyclic nucleotide analogs (PMEApp, HPMPApp and PMEDAPpp) in pol ε and pol ε* catalyzed reactions have shown that a proteolysis probably does not affect pol ε binding site for dNTPs. Both enzymes (pol ε and pol ε*) possess 3'-5'-exonuclease activity with different Km for 3'-OH end of template poly dA-oligo dT18 (1.6 μmol/l and 0.36 μmol/l, respectively).

scholarly journals THE Ah PHENOTYPE. SURVEY OF FORTY-EIGHT RAT STRAINS AND TWENTY INBRED MOUSE STRAINS

Genetics ◽  
1982 ◽  
Vol 100 (1) ◽  
pp. 79-87
Author(s):  
Daniel W Nebert ◽  
Nancy M Jensen ◽  
Hisashi Shinozuka ◽  
Heinz W Kunz ◽  
Thomas J Gill
Keyword(s):  
Specific Activity ◽  
Inbred Mouse ◽  
Inbred Mouse Strain ◽  
Mouse Strains ◽  
Ah Receptor ◽  
Inbred Mouse Strains ◽  
Sprague Dawley ◽  
Rat Strains ◽  
Specific Activities ◽  
Inbred Rat

ABSTRACT Forty-four inbred and four randombred rat strains and 20 inbred mouse strains were examined for their Ah phenotype by determining the induction of liver microsomal aryl hydrocarbon (benzo[a]pyrene) hydroxylase activity (EC 1.14.14.1) by intraperitoneal treatment with either β-naphthoflavone or 3-methylcholanthrene. All 48 rat strains were found to be Ah-responsive. The maximally induced hydroxylase specific activities of the ALB/Pit, MNR/Pit, MR/Pit, SHR/Pit, and Sprague-Dawley strains were of the same order of magnitude as the basal hydroxylase specific activities of the ACI/Pit, F344/Pit, OKA/Pit, and MNR/N strains. Six of the 20 mouse strains were Ah-nonresponsive (i.e. lacking the normal induction response and presumably lacking detectable amounts of the Ah receptor). The basal hydroxylase specific activities of the BDL/N, NFS/N, STAR/N, and ST/JN mouse strains were more than twice as high as the maximally induced hydroxylase specific activity of the CBA/HT strain.——To date, 24 Ah-nonresponsive mouse strains have been identified, out of a total of 68 known to have been characterized. The reasons for not finding a single Ah-nonresponsive inbred rat strain—as compared with about one Ah-nonresponsive inbred mouse strain found for every three examined—remain unknown.

Enzymatic condensation of dopamine and acetaldehyde: a salsolinol synthase from rat brain

Biologia ◽  
2011 ◽  
Vol 66 (6) ◽  
Author(s):  
Xuechai Chen ◽  
Abida Arshad ◽  
Hong Qing ◽  
Rui Wang ◽  
Jianqing Lu ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Substantia Nigra ◽  
Human Subjects ◽  
Enzymatic Reaction ◽  
Brain Regions ◽  
Activity Detection ◽  
Reaction Products ◽  
Sprague Dawley ◽  
Sprague Dawley Rats ◽  
Salsolinol Synthase

AbstractSalsolinol (1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline; Sal) is structurally similar to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, which is supposed to have a role in the development of Parkinson-like syndrome in both human and non-human subjects. In the human brain, the amount of (R)-enantiomer of Sal is much higher than (S)-enantiomer, suggesting that a putative enzyme may participate in the synthesis of (R)-salsolinol, called (R)-salsolinol synthase. In this study, the (R)-salsolinol synthase activity in the condensation of dopamine and acetaldehyde was investigated in the crude extracts from the brains of Sprague Dawley rats. Identification of the enzymatic reaction products and enzyme activity detection were achieved by HPLC-electrochemical detection. The discovery of this enzyme activity in rat’s brain indicates the natural existence of (R)-salsolinol synthase in the brains of humans and rats, and it is distributed in most brain regions of rat with higher activity in soluble proteins extracted from striatum and substantia nigra.

Effects of Echinomycin on PCNA-Dependent Follicular Development in PMSG-Induced Sprague-Dawley Rats

2014 ◽  
Vol 998-999 ◽  
pp. 228-232
Author(s):  
Zheng Hong Zhang ◽  
Fan Wang ◽  
Yan Qing Wu ◽  
Zong Hao Tang ◽  
Qing Qiang Lin ◽  
...  
Keyword(s):  
Hypoxia Inducible Factor ◽  
Follicular Development ◽  
Antral Follicle ◽  
Binding Activity ◽  
Nuclear Antigen ◽  
Sprague Dawley ◽  
Dna Binding Activity ◽  
Molecule Inhibitor ◽  
Protein Expressions ◽  
Serum Gonadotropin

Echinomycin (Ech) is a small-molecule inhibitor of hypoxia-inducible factor-1 DNA-binding activity, which plays a crucial role in the regulation of ovarian functions in mammals. The present study was designed to test the hypothesis that hypoxia-inducible factor (HIF)-1alpha-mediated proliferation cell nuclear antigen (PCNA) expressions contributed to the follicular development in the rat ovary primed by pregnant mare serum gonadotropin (PMSG). Through the histological examination, the decrease of growing and antral follicle numbers was found after Ech treatment both in control and PMSG treated groups. And then PCNA mRNA and protein expressions were found to significantly increase in the ovaries treated with PMSG, and the similar changes were found in HIF-1alpha mRNA and protein expressions, indicating PMSG-induced follicular development may be through HIF-1alpha/PCNA signaling. Furthermore, PCNA expression was found to significantly decrease in the ovaries after Ech treatment, while HIF-1alpha mRNA and protein expression was no obviously changes. Further analysis found the changes of PCNA expression were consistent with HIF-1 activity in the ovaries, further suggesting the regulatory roles in the follicular development. Taken together, these results demonstrated this HIF-1alpha-mediated PCNA expression is one of the important mechanisms regulating the ovarian follicular development in mammals. Keywords: HIF-1alpha; PCNA; echinomycin; HIF prolyl hyodroxylase acitvity; follicular development

Detection and characterization of DNA polymerase activity in Toxoplasma gondii

Parasitology ◽  
1993 ◽  
Vol 107 (2) ◽  
pp. 135-139 ◽  
Author(s):  
A. Makioka ◽  
B. Stavros ◽  
J. T. Ellis ◽  
A. M. Johnson
Keyword(s):  
Toxoplasma Gondii ◽  
Dna Polymerase ◽  
Dna Polymerases ◽  
Sedimentation Coefficient ◽  
Polymerase Activity ◽  
Magnesium Ions ◽  
Dna Polymerase Β ◽  
Human Dna ◽  
Approximate Molecular Weight

SUMMARYA DNA polymerase activity has been detected and characterized in crude extracts from tachzoites of Toxoplasma gondii. The enzyme has a sedimentation coefficient of 6·4 S, corresponding to an approximate molecular weight of 150000 assuming a globular shape. Like mammalian DNA polymerase α, the DNA polymerase of T. gondii was sensitive to N-ethylmaleimide and inhibited by high ionic strength. However, the enzyme activity was not inhibited by aphidicolin which is an inhibitor of mammalian DNA polymerases α, δ and ε and also cytosine-β-D-arabinofuranoside-5′-triphosphate which is an inhibitor of α polymerase. The activity was inhibited by 2′,3′-dideoxythymidine-5′-triphosphate which is an inhibitor of mammalian DNA polymerase β and γ. Magnesium ions (Mg2+) were absolutely required for activity and its optimal concentration was 6 mM. The optimum potassium (K+) concentration was 50 mM and a higher concentration of K+ markedly inhibited the activity. Activity was optimal at pH 8. Monoclonal antibodies against human DNA polymerase did not bind to DNA polymerase of T. gondii. Thus the T. gondii enzyme differs from the human enzymes and may be a useful target for the design of toxoplasmacidal drugs.

scholarly journals Exploration of anti-breast cancer effects of Terminalia chebula extract on DMBA-induced mammary carcinoma in Sprague Dawley rats

2020 ◽  
Vol 6 (1) ◽  
Author(s):  
Deena Priscilla Henry ◽  
Jasmine Ranjan ◽  
Rajesh Kumar Murugan ◽  
Annapoorani Sivanantham ◽  
Manikandan Alagumuthu
Keyword(s):  
Breast Cancer ◽  
Plant Extract ◽  
Mammary Carcinoma ◽  
Future Perspective ◽  
Terminalia Chebula ◽  
Sprague Dawley ◽  
Analytical Technique ◽  
Isolation And Characterization ◽  
Sprague Dawley Rats ◽  
Mcf 7

Abstract Background Plant extracts are effectively acting as the natural medicinal cocktail, non-side effective, efficacious, and freely available. The present study aimed to unveil the pharmacological and medicinal effects of Terminalia chebula plant extract in 7,12-dimethylbenzanthracene (DMBA)-induced mammary carcinoma in Sprague Dawley rats. The plant extract obtained was subjected to in vivo antioxidant and anticancer studies in various concentrations after an analytical technique such as FTIR, GCMS, and HPLC-based chemo-profiling in Sprague Dawley rats. Results Apart from the antiproliferative effect on breast cancer cell line (MCF-7) and normal breast epithelial cells (MCF-10a), we have measured the changes in body weight, along with other tumor parameters such as tumor volume, tumor incidence, tumor weight, tumor burden, serum biochemical parameters, and histopathological findings of breast tissue. As the oxidative stress further enhances the development of cancer, the antioxidant property of the plant extract demonstrates its use against cancer treatment. One hundred fifty milligrams per milliliter (IC50 250 μg/mL) concentration of the ethanolic extract was vital for the proliferation of MCF-7 cell lines (Fig. 7a). Meanwhile, 300 μg/mL (IC50 150 μg/mL) was an effective dose to attain a maximum HDAC inhibition of 78%. Also, the normal liver and kidney functioning revealed the non-toxicity nature of the plant. Conclusion Terminalia chebula could be one of the effective naturally obtained anti-breast cancer medications. Isolation and characterization of individual bioactive compounds of T. chebula would be the future perspective.

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