The Effect of Urea on the Structure of Nuclear Ribonucleoprotein Particles from Rat Liver

Ribonucleoprotein particles were isolated in the presence of cytosolic RNase inhibitor from rat liver nuclei and exposed to increasing concentrations of urea. The average sedimentation coefficient of 90 S in a sucrose gradient decreased slightly with urea concentrations of up to 2 M. At 5 M urea the particles sedimented at about 20 S. The protein‐to‐RNA ratio decreased from 4 to 1.4 after treatment with 5 M urea. The proteins which are preferentially detached from the ribonucleoprotein complex are in the molecular weight range of 30000–45000. In addition, small nuclear RNAs (snRNA) were dissociated from the ribonucleoprotein particles by high urea concentrations (> 2M). From the fact that snRNAs are dissociated from the ribonucleoprotein complex by high urea concentrations and not dissociated by high NaCl concentrations even after extensive proteolytic digestion, it is concluded that hydrogen bonds are involved in their binding to heterogeneous nuclear RNA (hnRNA). The change in the RNA conformation after addition of increasing concentrations of urea was studied by circular dichroism measurements of ribonucleoprotein particles and protein‐free particle RNA. In the presence of 5 M urea the molar ellipticities at 264 nm decreased from 15800 to 9400, 21000 to 19700, 19500 to 16600 and from 20900 to 16800 deg × cm 2 × dmol −1 in the case of ribonucleoprotein particles, protein‐free particle RNA, hnRNA and snRNA respectively. From the results of the circular dichroism measurements it is concluded that the high‐molecular‐weight proteins, which are not dissociated from the particles, are responsible for a conformational restraint on hnRNA. snRNA, hnRNA, as well as the physiological mixture of both, exhibit the same gradual increase in absorbance with temperature. In the presence of 0.1 M NaCl melting temperatures of about 43°C were obtained in all three cases. The gradual nature of the thermal transition is due to the shortness and heterogeneity of the helical regions. The average helix length is about 5 base pairs long.