Fig. 4.

Large-scale purification and crystallization of the P. pastoris RNA polymerase II. (a) Large-scale purification of RNAP II. Protein fraction of each purification step was separated on a 10–20 % SDS polyacrylamide gel and visualized by CBB staining. M, molecular weight markers. The molecular weights (kDa) are shown on the left. Lane 1, total cell lysate; lane 2, elution from Q Sepharose; lane 3, elution from the TAP purification; and lane 4, elution from Resource Q. Aro2p is marked with an asterisk in lane 3. In lane 4, the THF-tagged Rpb2 subunit and its co-purified subunits are marked with an open triangle and filled triangles, respectively. Two small subunits, Rpb10 (8.5 kDa) and Rpb12 (7.9 kDa), seemed to co-migrate at the position marked with #. This figure is composed of multiple gel images. (b) Single crystals of P. pastoris RNAP II. (c) X-ray diffraction image of an RNAP II crystal