Structural transitions mediating transcription initiation by T7 RNA polymerase.
During transcription initiation, RNA polymerases appear to retain promoter interactions while transcribing short RNAs that are frequently released from the complex. Upon transition to elongation, the polymerase releases promoter and forms a stable elongation complex. Little is known about the changes in polymerase conformation or polymerase:DNA interactions that occur during this process. To characterize the transitions that occur in the T7 RNA polymerase transcription complex during initiation, we prepared enzymes with Fe-BABE conjugated at 11 different positions. Addition of H(2)O(2) to transcription complexes prepared with these enzymes led to nucleic acid strand scission near the conjugate. Changes in the cleavage sites revealed a series of conformational changes and rearrangements of protein:nucleic acid contacts that mediate progression through the initiation reaction.
Mukherjee
Srabani
S
Department of Biochemistry, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, TX 78229, USA.
Brieba
Luis G
LG
Sousa
Rui
R
D001665:Binding Sites; D003545:Cysteine / Q000737:chemistry / Q000235:genetics; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000378:metabolism; D004492:Edetic Acid / Q000031:analogs & derivatives* / Q000737:chemistry; D007502:Iron Chelating Agents / Q000737:chemistry; D018625:Microscopy, Atomic Force; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D009942:Organometallic Compounds / Q000737:chemistry; D011487:Protein Conformation; D012313:RNA / Q000378:metabolism; D013329:Structure-Activity Relationship; D024363:Transcription Initiation Site; D014158:Transcription, Genetic; D014764:Viral Proteins
D007502:Iron Chelating Agents; D009942:Organometallic Compounds; D014764:Viral Proteins; D012313:RNA; C042007:1-(4-bromoacetamidobenzyl)EDTA; D004247:DNA; D004492:Edetic Acid; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; D003545:Cysteine