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The purification from rat liver of a nuclease hydrolysing ribonucleic acid and deoxyribonucleic acid.

ABSTRACT: 1. The purification of a nuclease from rat-liver mitochondria is described. The mitochondria are rendered soluble by treatment with Triton X-100 and, after fractionation with ammonium sulphate and acetone, the active fraction is further purified by chromatography on DEAE-cellulose and Sephadex G-75 to give a purification of over 700-fold. 2. The purified enzyme was only very slightly contaminated with deoxyribonuclease II, phosphodiesterase and phosphomonoesterase. The individual activities of these enzymes did not exceed 0.1% of the activity of the liver nuclease. 3. The purified enzyme attacked RNA more rapidly than denatured DNA and hydrolysed native DNA more slowly than denatured DNA. 4. There is some evidence to suggest that the nucleolytic activity of the purified preparation towards native DNA, denatured DNA and RNA is associated with a single protein. 5. The enzyme is relatively labile but is stabilized in the presence of 20% (w/v) glycerol or 10mm-2-mercaptoethanol.

SUBMITTER: Curtis PJ

PROVIDER: S-EPMC1264923 | biostudies-other | 1966 Mar

REPOSITORIES: biostudies-other

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Project description:1. The stimulations of DNA-dependent RNA polymerase in isolated rat-liver nuclei by thyroid hormone, human growth hormone and testosterone are compared. 2. Single or multiple administrations of growth-promoting doses of tri-iodo-l-thyronine, human growth hormone and testosterone stimulate the Mg(2+)-activated RNA-polymerase reaction in nuclei from thyroidectomized, hypophysectomized and castrated rats respectively. The magnitude of stimulation was proportional to the degree of enhancement of liver growth by each hormone. After a single injection, the latent period preceding the stimulation was 1, 2 and 10hr. for growth hormone, testosterone and tri-iodothyronine respectively. The time-course of stimulation of enzyme activity and the synthesis of rapidly labelled nuclear RNA in vivo were also different for each hormone. 3. Growth hormone administration failed to stimulate the Mn(2+)/ammonium sulphate-activated RNA-polymerase reaction. Thyroid hormone and testosterone, however, stimulated it but the effect was less pronounced and occurred several hours later than that observed for the Mg(2+)-activated RNA-polymerase reaction. 4. In combination experiments, hypophysectomized or the thyroidectomized rats were given growth hormone or tri-iodothyronine in a single or repeated doses at levels that produced the maximum stimulation of Mg(2+)-activated RNA-polymerase activity. Taking into account the different latent period for each hormone, a single administration of the second hormone caused an additional stimulation of the enzyme activity. Similar additive effects were observed in thyroidectomized-castrated rats after treatment with tri-iodothyronine and testosterone. The magnitude of the additional stimulation caused by the administration of the second hormone was compatible with the capacity of that hormone to promote liver growth in rats deprived of it. 5. It is concluded that, although these hormones have some similar effects, the regulation of nuclear RNA synthesis may be mediated via different routes for each hormone.

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The purification from rat liver of a nuclease hydrolysing ribonucleic acid and deoxyribonucleic acid.

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