Đang chuẩn bị nút TẢI XUỐNG, xin hãy chờ
Tải xuống
BacterialRNApolymerase (RNAP) is the central enzyme of gene expression that is responsible for the synthesis of all types of cellular RNAs. The process of transcription is accompanied by complex structural rearrangements of RNAP. Despite the recent progress in structural studies of RNAP, detailed mechanisms of conformational changes of RNAP that occur at different stages of transcription remain unknown. The goal of this workwas to obtain novel ligands to RNAP which would target different epitopes of the enzyme and serve as specific probes to study the mech-anism of transcription and conformational flexibility of RNAP | Eur. J. Biochem. 271 4921-4931 2004 FEBS 2004 doi 10.1111 j.1432-1033.2004.04461.x Aptamers to Escherichia coli core RNA polymerase that sense its interaction with rifampicin r-subunit and GreB Andrey Kulbachinskiy1 2 Andrey Feklistov2 3 Igor Krasheninnikov3 Alex Goldfarb1 and Vadim Nikiforov1 2 1Public Health Research Institute Newark New Jersey USA 2Institute of Molecular Genetics Moscow Russia 3Department of Molecular Biology Moscow State University Moscow Russia Bacterial RNA polymerase RNAP is the central enzyme of gene expression that is responsible for the synthesis of all types of cellular RNAs. The process of transcription is accompanied by complex structural rearrangements of RNAP. Despite the recent progress in structural studies of RNAP detailed mechanisms of conformational changes of RNAP that occur at different stages of transcription remain unknown. The goal of this work was to obtain novel ligands to RNAP which would target different epitopes of the enzyme and serve as specific probes to study the mechanism of transcription and conformational flexibility of RNAP. Using in vitro selection methods we obtained 13 classes of ssDNA aptamers against Escherichia coli core RNAP. The minimal nucleic acid scaffold an oligonucleotide construct imitating DNA and RNA in elongation complex rifampicin and the ơ70-subunit inhibited binding of the aptamers to RNAP core but did not affect the dissociation rate of preformed RNAP-aptamer complexes. We argue that these ligands sterically block access of the aptamers to their binding sites within the main RNAP channel. In contrast transcript cleavage factor GreB increased the rate of dissociation of preformed RNAP-aptamer complexes. This suggested that GreB that binds RNAP outside the main channel actively disrupts RNAP-aptamer complexes by inducing conformational changes in the channel. We propose that the aptamers obtained in this work will be useful for studying the interactions of RNAP with various ligands and .
