Calf thymus DNA-Topoisomerase I activity was found to be altered by changing in phosphorylation: it was completely inhibited upon dephosphorylation by alkaline phosphatase, but incubation with N II protein kinase and ATP restored the relaxation activity to a level higher than that observed prior to dephosphorylation. The calf thymus Topoisomerase I-mediated DNA cleavage, induced by camptothecin, also proved to be inhibited by dephosphorylation, which, apparently, stabilizes the initial enzyme-substrate complex. We conclude that: the native protein is partially phosphorylated, the phosphorylation involvement is essential for the activity expression and also for DNA-protein interaction, changes in the degree of phosphorylation might be involved in the regulation of DNA processing; that evokes some properties of chromatinic peptide models, which bind DNA only when phosphorylated and leads to the assumption that they represent the minimum functional substrate for N II protein kinase.

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Biotin deficient rat liver histones showed decreased phosphorylation and methylation, and increased acetylation rates as compared to normal rat liver histones: these alterations may be related to the observed lower stability of the interactions between histones and DNA. The modifications of the metabolic process might be the consequence of an alteration of the synthesis of the enzymes involved in histone phosphorylation, acetylation and methylation mechanisms and are presumably related to a biotin effect upon the synthesis of RNA and proteins.

Reactivity and chemical properties of calf thymus topoisomerase I have been investigated with respect to enzyme ability to relax supercoiled DNA. The relaxation rate has been analyzed at optimum and relatively high salt concentration. Catalysis is processive at optimum salt concentration and distributive at a higher one; camptothecin decreases the initial rate of reaction in both salt conditions, but more so at the higher one. We conclude that: 1. calf thymus topoisomerase I requires, for its maximum reactivity, specific and characteristic reaction conditions; 2. salt concentration affects DNA processing, indeed influencing the initial rate of DNA relaxation and directly reflecting the salt-dependence for the enzyme-duplex DNA binding; 3. topoisomerase I, from various sources, maybe individually responds to alteration of assay parameters such as pH, Mg++ and NaCl concentrations, indicating that individual criteria could be responsible for the catalytic activity optimum.

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