Large scale isolation of RNA polymerase I of Tetrahymena pyriformis and the identification of polymerase-associated polypeptides.

A procedure has been developed for a large scale and rapid isolation of RNA polymerase I (EC 2.7.7.6) of Tetrahymena pyriformis. The enzyme is precipitated from the cell homogenate by Polymin P, extracted from the sediment and separated from RNA polymerase II by a treatment with phosphocellulose. The further purification procedure involves sedimentation in glycerol gradients and chromatography on heparin-Sepharose and DEAE-Sephadex. The last step achieved the separation of RNA polymerase I from RNA polymerase III. On the basis of different criteria RNA polymerase I is assumed to consist of two large subunits of 180 and 118 kDa and nine subunits smaller than 50 kDa. Additional polypeptides have been identified which are associated with RNA polymerase I but are not found in integral stoichiometric amounts. Except for certain minor differences RNA polymerase I purified from the cell homogenate shows the same structure as the enzyme obtained from isolated macronuclei (Mueller et al., 1985).