Inactivation of the Bacterial RNA Polymerase Due to Acquisition of Secondary Structure by the omega Subunit

Sarkar, Paramita and Sardesai, Abhijit A and Murakami, Katsuhiko S and Chatterji, Dipankar (2013) Inactivation of the Bacterial RNA Polymerase Due to Acquisition of Secondary Structure by the omega Subunit. In: JOURNAL OF BIOLOGICAL CHEMISTRY, 288 (35). pp. 25076-25087.

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Abstract

The widely conserved omega subunit encoded by rpoZ is the smallest subunit of Escherichia coli RNA polymerase (RNAP) but is dispensable for bacterial growth. Function of omega is known to be substituted by GroEL in omega-null strain, which thus does not exhibit a discernable phenotype. In this work, we report isolation of omega variants whose expression in vivo leads to a dominant lethal phenotype. Studies show that in contrast to omega, which is largely unstructured, omega mutants display substantial acquisition of secondary structure. By detailed study with one of the mutants, omega(6) bearing N60D substitution, the mechanism of lethality has been deciphered. Biochemical analysis reveals that omega(6) binds to beta ` subunit in vitro with greater affinity than that of omega. The reconstituted RNAP holoenzyme in the presence of omega(6) in vitro is defective in transcription initiation. Formation of a faulty RNAP in the presence of mutant omega results in death of the cell. Furthermore, lethality of omega(6) is relieved in cells expressing the rpoC2112 allele encoding beta ` (2112), a variant beta ` bearing Y457S substitution, immediately adjacent to the beta ` catalytic center. Our results suggest that the enhanced omega(6)-beta ` interaction may perturb the plasticity of the RNAP active center, implicating a role for omega and its flexible state.

Item Type:	Journal Article
Publication:	JOURNAL OF BIOLOGICAL CHEMISTRY
Publisher:	AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Additional Information:	Copyright for this article belongs to the AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, USA
Department/Centre:	Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited:	10 Mar 2014 06:35
Last Modified:	10 Mar 2014 06:35
URI:	http://eprints.iisc.ac.in/id/eprint/48519

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