Structural basis for the transition from initiation to elongation transcription in T7 RNA polymerase.
To make messenger RNA transcripts, bacteriophage T7 RNA polymerase (T7 RNAP) undergoes a transition from an initiation phase, which only makes short RNA fragments, to a stable elongation phase. We have determined at 2.1 angstrom resolution the crystal structure of a T7 RNAP elongation complex with 30 base pairs of duplex DNA containing a "transcription bubble" interacting with a 17-nucleotide RNA transcript. The transition from an initiation to an elongation complex is accompanied by a major refolding of the amino-terminal 300 residues. This results in loss of the promoter binding site, facilitating promoter clearance, and creates a tunnel that surrounds the RNA transcript after it peels off a seven-base pair heteroduplex. Formation of the exit tunnel explains the enhanced processivity of the elongation complex. Downstream duplex DNA binds to the fingers domain, and its orientation relative to upstream DNA in the initiation complex implies an unwinding that could facilitate formation of the open promoter complex.
Yin
Y Whitney
YW
Department of Molecular Biophysics and Biochemistry, Yale University, 266 Whitney Avenue, New Haven, CT 06520-8114, USA.
Steitz
Thomas A
TA
D017123:Bacteriophage T7 / Q000201:enzymology; D001665:Binding Sites; D003460:Crystallization; D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000378:metabolism; D009692:Nucleic Acid Heteroduplexes; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D012319:RNA Polymerase II / Q000737:chemistry; D012333:RNA, Messenger / Q000737:chemistry* / Q000378:metabolism; D019914:Taq Polymerase / Q000737:chemistry; D013698:Templates, Genetic; D024363:Transcription Initiation Site; D014158:Transcription, Genetic; D014764:Viral Proteins
D009692:Nucleic Acid Heteroduplexes; D021122:Protein Subunits; D012333:RNA, Messenger; D014764:Viral Proteins; D004247:DNA; D012319:RNA Polymerase II; D019914:Taq Polymerase; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; D009238:N-Acetylmuramoyl-L-alanine Amidase