Specific thyroxine nuclear receptors: DEAE-sephadex chromatography of rat liver nuclear extract

Specific binding of thyroxine to protein fraction obtained by DEAE-Sephadex chromatography of dialyzed (0.4 mol 1-1 KCl) nuclear extract of purified rat liver nuclei was studied. A procedure for several-fold purification of thyroxine nuclear receptor included isolation of purified rat liver nuclei (by centrifugation through 1.7 mol 1-1 sucrose), extraction, dialysis, Sephadex G-100 and DEAE-Sephadex A-50 chromatography. It was found that a protein peak eluting at 0.05 mol 1-1 NaCl does not show any fraction specifically binding thyroxine. However, specific binding of nuclear receptor for thyroxine was found in a peak eluting at 0.20 mol 1-1 NaCl, the capacity being about 600 fmol mg-1 protein. Moreover, total binding capacity of intact rat liver homogenate, nuclei, non-dialyzed and dialyzed (0.4 mol 1-1 KCl) nuclear extract, Sephadex G-100 and DEAE-Sephadex A-50 chromatography protein fractions for thyroxine was tested. A marked increase (about 20 fold) in the total thyroxine binding expressed in fmol mg-1 of protein in DEAE-Sephadex A-50 eluate was found as compared to whole nuclei, nuclear extract or Sephadex G-100 chromatography fractions. Partial fractionation of rat liver nuclear extract combined with thyroxine binding studies demonstrated that rat liver nuclei contain a non-histone protein fraction--a receptor that is responsible for a specific binding of thyroxine.