On the determination of the sedimentation coefficient of DNA in a preparative ultracentrifuge

1972 ◽  
Vol 37 (6) ◽  
pp. 1938-1942 ◽  
Author(s):  
J. Huťková ◽  
V. Drášil
Keyword(s):  
Sedimentation Coefficient

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).

Strahlenempfindlichkeit von Bakteriophagen-DNS /Radiation sensitivity of Bacteriophage DNA

1969 ◽  
Vol 24 (7) ◽  
pp. 885-893 ◽  
Author(s):  
Thérèse Coquerelle ◽  
Leuthold Bohne ◽  
Ulrich Hagen ◽  
Jürgen Merkwitz
Keyword(s):  
Molecular Weight ◽  
Average Molecular Weight ◽  
Linear Part ◽  
Sedimentation Coefficient ◽  
Radiation Sensitivity ◽  
Quadratic Term ◽  
Molecular Weight Distributions ◽  
Γ Rays ◽  
Higher Doses

DNA isolated from Coli bacteriophage T1 was irradiated in 0.165 ᴍ NaCl with γ-rays. The molecular weight was determined by measurement of the sedimentation coefficient and viscosity. An analysis of the boundary permits the determination of the sedimentation distribution. The distribution of sedimentation coefficients obtained at various DNA concentrations were extrapolated to zero concentration and were transformed into molecular weight distributions. These were used to calculate the weight average molecular weight Mw and the number average molecular weight Mn.The molecular weight of T1-DNA was found to be 32· 106. After irradiation at a concentration of 200 μg/ml, double breaks as well as intermolecular crosslinks could be determined. The number of double breaks showed a rise with dose that can best be described as composed of a linear and a quadratic term. At low doses the crosslinks increase linearly, the rate being approximately half of that for the linear part of the double breaks. After higher doses, where most of the molecules are degraded, apparently no additional crosslinks are produced. No crosslinks were seen in DNA degraded by DNase. The influence of the DNA concentration on the degradation and the formation of crosslinks is discussed.

Anreicherung der Leber-N-Acetyltransferase durch präparative Polyacrylamidgel-Elektrophorese und Bestimmung des Molekulargewichts /Purification of Human Liver Serotonin-/Isoniazid-N-Acetyltransferase by Preparative Polyacrylamidgel-Electrophoresis and Determination of Molecular Weight

1974 ◽  
Vol 29 (11-12) ◽  
pp. 661-666 ◽  
Author(s):  
Ernst Schulte ◽  
Werner Schloot ◽  
H. Werner Goedde
Keyword(s):  
Molecular Weight ◽  
Column Chromatography ◽  
Human Liver ◽  
Sedimentation Coefficient ◽  
Disc Electrophoresis

Abstract Human liver N-acetyltransferase was purified by means of column chromatography on Sephadex G-100 (18 fold; yield: 60-70%) and preparative disc electrophoresis (40 fold; yield: 95 - 100%). A separation of the hypothetical “serotonin-N-acetyltransferase” and “INH-N-acetyltransferase” on the basis of a possible difference of the charge of the proteins could not be achieved (disc electrophoresis). Thus the assumption is confirmed that INH and Serotonin are substrates of one enzyme. The activity of N-acetyltransferase can be stabilized by adding thioglycolate, 2-mercaptoethanol, and cysteine. In some cases an activation of 100 to 200% could be observed. A molecular weight of about 26 500 was determined by the method of column chromatography on Sephadex as well as by calculating the sedimentation coefficient. Enzymes prepared by different procedures showed similar Kм-values.

scholarly journals Haemoglobin from the tadpole shrimp, Lepidurus apus lubbocki Characterization of the molecule and determination of the number of polypeptide chains

1979 ◽  
Vol 183 (2) ◽  
pp. 325-330 ◽  
Author(s):  
E Ilan ◽  
E Daniel
Keyword(s):  
Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Sedimentation Coefficient ◽  
Sedimentation Equilibrium ◽  
Guanidinium Chloride ◽  
Tadpole Shrimp ◽  
Polypeptide Chains

Haemoglobin from the tadpole shrimp, Lepidurus apus lubbocki, was found to have a sedimentation coefficient (s020,w) of 19.3 +/- 0.2 S and a molecular weight, as determined by sedimentation equilibrium, of 798000 +/- 20000. The amino acid composition showed the lack of cysteine and cystine residues. A haem content of 3.55 +/- 0.03% was determined, corresponding to a minimal mol.wt. of 17400 +/- 200. The pH-independence in the range pH 5-11 of the sedimentation coefficient indicates a relatively high stability of the native molecule. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis gave one band with mobility corresponding to a mol.wt. of 34000 +/- 1500. The molecular weight of the polypeptide chain was determined to be 32800 +/- 800 by sedimentation equilibrium in 6 M-guanidinium chloride and 0.1 M-2-mercaptoethanol. The findings indicate that Lepidurus haemoglobin is composed of 24 identical polypeptide chains, carrying two haem groups each.

CONGENITAL GOITRE IN SHEEP: ISOLATION OF THE IODOPROTEINS WHICH REPLACE THYROGLOBULIN

1976 ◽  
Vol 71 (2) ◽  
pp. 179-192 ◽  
Author(s):  
C. E. DOLLING ◽  
B. F. GOOD
Keyword(s):  
Molecular Weight ◽  
Thyroid Gland ◽  
Sucrose Gradient ◽  
Sedimentation Coefficient ◽  
Agarose Gel ◽  
Merino Sheep ◽  
Thyroid Glands ◽  
Sucrose Gradient Ultracentrifugation ◽  
Australian Merino

SUMMARY Immunological and chromatographic studies of proteins from the congenital goitre of South Australian Merino sheep revealed that normal thyroglobulin is absent from the thyroid glands of these sheep. However, a thyroglobulin-immunoreactive iodoprotein was isolated by affinity chromatography on agarose gel to which antibody against thyroglobulin had been covalently bound. Sucrose-gradient ultracentrifugation indicated that this iodoprotein had a sedimentation coefficient of 8S and a molecular weight of approximately 175000. This iodoprotein is therefore about one quarter the size of normal thyroglobulin (19S; 660000) and is similar in size to the subunit of thyroglobulin (3-8S; 165000) although this has usually been described as non-iodinated except when derived by reductive fission. In addition the goitre extract contained iodoproteins which had the immunological properties of serum albumin and immunoglobulin G. Determination of the iodine and iodoamino acid content of the hydrolysed iodoproteins revealed that they contained iodothyronines which were able to contribute to the production of thyroid hormones although the total iodothyronine content of the goitrous gland was less than that of the normal sheep thyroid gland.

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