Two-dimensional and epitaxial crystallization of a mutant form of yeast RNA polymerase II*1

Abstract

A mutant form of yeast RNA polymerase II that lacks the fourth and seventh largest subunits, referred to as pol II Δ 4/7, crystallized on positively charged lipid layers. Both single-layered (two-dimensional) crystals and several multi-layered crystal forms were obtained. The two-dimensional crystals, preserved in negative stain, diffracted strongly to about 120 Å−1 and more weakly to 1/13 Å−1 resolution. A projection map computed from averaged Fourier transforms revealed four pol II Δ 4/7 complexes per unit cell and further revealed a cleft on the surface of the complex similar to that previously observed in the structure of Escherichia coli RNA polymerase. One of the multi-layered crystal forms, preserved in negative stain, diffracted strongly beyond 1/15 Å−1 resolution. Coherent diffraction from the multi-layered crystal is indicative of protein-protein interactions between layers and ordering in the third dimension.