PURIFICATION AND PROPERTIES OF ESTROGEN-RESPONSIVE CYTOPLASMIC THYMIDINE KINASE FROM HUMAN-BREAST CANCER

  • 1 January 1981
    • journal article
    • research article
    • Vol. 41  (2) , 604-610
Abstract
The effect of 17.beta.-estradiol on cytoplasmic thymidine kinase activity was studied in MCF-7, a human breast cancer cell line in culture which responds to estrogens with an increase in the rate of growth. Levels of 17.beta.-estradiol which maximally stimulate [3H]thymidine incorporation into DNA also maximally stimulate thymidine kinase activity. The Vmax for thymidine increased while the Km was not affected by estrogen stimulation when performed on nonpurified enzyme. Tamoxifen, an antiestrogen, decreased the specific activity of the enzyme. To further study its hormonal regulation, cytoplasmic thymidine kinase was purified > 2000-fold by affinity column chromatography. The purified preparation migrated in 1 band to a pl [isolectric point] of 8.5 on an isoelectric focusing gel. The purified thymidine kinase was further characterized by examining its MW, pH optimum, heat stability, use of phosphate donors, inhibition by nucleotides and the effect of pyrimidine nucleoside analogs.