Formation and crystallization of yeast RNA polymerase II elongation complexes.
Minimal templates were devised for the efficient generation of yeast RNA polymerase II transcription elongation complexes. A 33-base pair DNA with a 15-residue dC tail at one 3'-end supported the formation of a complex containing the polymerase paused at nucleotide 11 of the duplex region and an RNA of 14-16 residues. The same template could yield an arrested complex with the enzyme at nucleotide 13-15 and RNA of 15-17 residues. These complexes were stable for at least a week under various conditions and could be resolved by gel electrophoresis or purified by ion exchange chromatography. The purified paused complex formed crystals capable of x-ray diffraction to 3.5 A resolution. The complex remained active in the crystal and, in the presence of nucleoside triphosphates, could efficiently extend the transcript in situ.
Gnatt
A
A
Department of Structural Biology, Stanford University School of Medicine, Stanford California, 94305, USA.
Fu
J
J
Kornberg
R D
RD
D001483:Base Sequence; D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D010441:Peptide Chain Elongation, Translational; D011487:Protein Conformation; D012319:RNA Polymerase II / Q000737:chemistry* / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D013698:Templates, Genetic; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism; D035165:Transcription Factors, General; D014158:Transcription, Genetic; D035602:Transcriptional Elongation Factors
D014157:Transcription Factors; D035165:Transcription Factors, General; D035602:Transcriptional Elongation Factors; C018648:transcription factor S-II; D004247:DNA; D012319:RNA Polymerase II