Interaction of rifamycin with bacterial RNA polymerase.

Wehrli, W.; Knüsel, F.; Schmid, Karl; Staehelin, Matthys

doi:10.1073/pnas.61.2.667

Cited by 281 publications

(126 citation statements)

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“…Wehrli and Staehelin have shown that even in the presence of 0.5 M NH,C1 [6] or 1 M KC1 [2] rifampicin forms a complex with monomeric RNA polymerase which is stable enough to be isolated by gel filtration. and 0.04 mM dithiothreitol for 15 min a t 0 "C (total volume 0.97 ml).…”

Section: Influence Of Freezing and Thawing At High Ionic Strengthmentioning

confidence: 99%

“…Several observations point to the binding of the antibiotic to RNA polymerase as the immediate cause of inhibition of RNA synthesis [2,3]. Previous studies have made it unlikely that rifamycins are bound to the enzyme by a covalent bond [3].…”

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confidence: 99%

“…Rifampicin, a potent inhibitor of bacterial RNA synthesis [I], forms very rapidly a stable complex with RNA polymerase from Escherichia coli [2,3] even a t low temperatures. Several observations point to the binding of the antibiotic to RNA polymerase as the immediate cause of inhibition of RNA synthesis [2,3].…”

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On the Binding of Rifampicin to the DNA‐Directed RNA Polymerase from Escherichia coli

Lill

Hartmann

1973

European Journal of Biochemistry

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The capacity of RNA polymerase to bind the antibiotic rifampicin is rather sensitive to conditions which lead to conformational changes in the enzyme. Repeated freezing and thawing in I M LiCl as well as treatment with guanidinium chloride destroys the binding affinity of the enzyme for the inhibitor. Similarly rifampicin does not associate with any of the isolated subunits a, /3 or /3' . Surprisingly, the reaction of an excess of mercurials with the enzyme leads only to a partial loss of binding capacity suggesting that -SH groups are not directly involved in the formation of the complex with rifampicin. During chain elongation very little rifampicin is bound to the enzyme. Simultaneously enzymatic activity is not affected by the inhibitor. RNA cannot protect the enzyme against the attachment of rifampicin although the binding capacity is reduced. Treatment of the complex of enzyme and antibiotic with RNA does not release rifampicin from the complex. These results support the hypothesis that the tight binding of rifampicin to RNA polymerase is the principal cause of the inhibitory action of the antibiotic. . Probably other forces such as hydrogen bonds and hydrophobic interactions between different parts of the antibiotic molecule and a specific region of the three-dimensional structure of the enzyme are responsible for the tight binding. Complex formation should then by rather sensitive to agents which induce conformational changes of the enzyme. Usually the geometrical structure of proteins is sensitive to variation of the ionic strength of the medium. This is also true for RNA polymerase from E. coli. At high ionic strength the enzyme exists as a monomer whereas a t lower ionic strength a dimer is formed in the presence of sigma factor [4,5] or a Dedicated to Professor Karl Winnacker on the occasion of his 70th birthday.Enzyme. DNA-dependent RNA polymerase or nucleosidetriphosphate: RNA nucleotidyltransferase (EC 2.7.7.6).Definition. A,,, unit, the quantity of material contained in 1 ml of a solution which has an absorbance of 1 a t 260 nm when measured in a 1-cm pathlength cell. polymeric aggregate if sigma is absent [5]. Surprisingly, however, the stoichiometry of the complex of rifampicin and the monomeric, dimeric [6] or polymeric [7] form of the enzyme, respectively is the same independent of the presence of sigma. Each form of complex contains about one molecule of antibiotic per monomeric unit of RNA polymerase. I n addition the stability of these complexes does not differ noticeably [7]. I n view of these results we were interested to learn more about conditions and agents which could lead to changes in the secondary, tertiary and quaternary structure of RNA polymerase with concomitant loss of the capacity to bind rifampicin. I n this context the binding of the antibiotic to isolated subunits of RNA polymerase has also been studied.This problem has some bearing on the well-known fact that rifampicin inhibits specifically the initiation of the RNA polymerase reaction prior to the addition of substrate...

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Section: Influence Of Freezing and Thawing At High Ionic Strengthmentioning

confidence: 99%

mentioning

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On the Binding of Rifampicin to the DNA‐Directed RNA Polymerase from Escherichia coli

Lill

Hartmann

1973

European Journal of Biochemistry

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“…2) produced by an actinobacterium Amycolatopsis mediterranei [4]. In general, rifamycins bind to the b-subunit of the RNA polymerase, and inhibits mRNA transcription [5,6]. Introduction of rifampicin in late 1960s led to considerable fall in the mortality rate due to TB and since then it has been used as the first line of drug.…”

Section: Introductionmentioning

confidence: 99%

Synthetic Biology in Action: Developing a Drug Against MDR-TB

et al. 2014

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The amalgamation of the research efforts of biologists, chemists and geneticists led by scientists at the Department of Zoology, University of Delhi has resulted in the development of a novel rifamycin derivative; 24-desmethylrifampicin, which is highly effective against multi-drug resistant (MDR) strains of Mycobacterium tuberculosis. The production of rifamycin analogue was facilitated by genetic-synthetic strategies that have opened an interdisciplinary route for the development of more such rifamycin analogues aiming at a better therapeutic potential. The results of this painstaking effort of nearly 25 years of a team of students and scientists led by Professor Rup Lal have been recently published in the Journal of Biological Chemistry (www.jbc.org/content/289/30/21142. long). This strategy can now find applications for developing newer rifamycin analogues that can be harnessed to overcome the problem of MDR, extensively drug resistant (XDR) and totally drug resistant (TDR) M. tuberculosis.

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“…Together with biochemical evidence (5,34) this indicates that streptolydigin inhibits RNA synthesis by interaction with the RNA polymerase rather than by binding to the template as does actinomycin D (27). In contrast to rifampin, which acts on initiation of transcription (36,41), streptolydigin has been shown to interfere with RNA chain elongation (5). Cassani et al (5) also showed stabilization of the RNA-enzyme-deoxyribonucleic acid (DNA) complex in vitro when using purified RNA polymerase and T4 DNA; the inhibition of RNA chain elongation is reversible in vitro (34).…”

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Reevaluation of the Mode of Action of Streptolydigin in Escherichia coli: Induction of Transcription Termination In Vivo

Meyenburg

Nielsen

Johnsen

et al. 1978

Antimicrob Agents Chemother

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Growth of the permeable strain AS19 of Escherichia coli B is more sensitive to the antibiotic streptolydigin than is in vitro ribonucleic acid (RNA) synthesis. The in vivo chain elongation rates of lacZ messenger RNA and ribosomal RNA are not affected at 1.5 x 10-6 M, a concentration that reduces the growth rate threefold. The synthesis of large proteins is inhibited preferentially, and a considerable fraction of the polypeptides synthesized is unstable. The synthesis of complete ,8-galactosidase is inhibited relative to the synthesis of short, unstable polypeptides, which include the first 60 to 70 amino acids of /3-galactosidase. The expression of the following polycistronic transcription units is strongly biased against promoter-distal genes: trp, deo, rpoBC, and rrn. The extent of polarity is proportional to the distance transcribed and to the streptolydigin concentration. Streptolydigin appears to destabilize active transcription complexes irreversibly irrespective of the type of transcript (messenger RNA, ribosomal RNA) and of transcription intensity. We suggest that streptolydigin leads to premature termination of transcription, resulting in release of incomplete transcripts and, thus, a decrease in overall messenger RNA concentration, which becomes limiting for protein synthesis, i.e., for growth.The antibiotic streptolydigin interferes with bacterial ribonucleic acid (RNA) synthesis in vivo and in vitro (32, 34). Mutations to streptolydigin resistance are located in the /B subunit of the RNA polymerase (13,16,32). Together with biochemical evidence (5, 34) this indicates that streptolydigin inhibits RNA synthesis by interaction with the RNA polymerase rather than by binding to the template as does actinomycin D (27). In contrast to rifampin, which acts on initiation of transcription (36, 41), streptolydigin has been shown to interfere with RNA chain elongation (5). Cassani et al. (5) also showed stabilization of the RNA-enzyme-deoxyribonucleic acid (DNA) complex in vitro when using purified RNA polymerase and T4 DNA; the inhibition of RNA chain elongation is reversible in vitro (34).In attempts to apply these in vitro results in vivo, we realized that the effects of streptlydigin

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Interaction of rifamycin with bacterial RNA polymerase.

Cited by 281 publications

References 12 publications

On the Binding of Rifampicin to the DNA‐Directed RNA Polymerase from Escherichia coli

On the Binding of Rifampicin to the DNA‐Directed RNA Polymerase from Escherichia coli

Synthetic Biology in Action: Developing a Drug Against MDR-TB

Reevaluation of the Mode of Action of Streptolydigin in Escherichia coli: Induction of Transcription Termination In Vivo

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