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      The moonlighting function of bacteriophage P4 capsid protein, Psu, as a transcription antiterminator

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          Abstract

          Psu, a 20-kD bacteriophage P4 capsid decorating protein moonlights as a transcription antiterminator of the Rho-dependent termination. Psu forms specific complex with E.coli Rho protein, and affects the latter's ATP-dependent translocase activity along the nascent RNA. It forms a unique knotted dimer to take a V-shaped structure. The C-terminal helix of Psu makes specific contacts with a disordered region of Rho, encompassing the residues 139–153. An energy minimized structural model of the Rho–Psu complex reveals that the V-shaped Psu dimer forms a lid over the central channel of the Rho hexamer. This configuration of Psu causes a mechanical impediment to the translocase activity of Rho. The knowledge of structural and mechanistic basis of inhibition of Rho action by Psu may help to design peptide inhibitors for the conserved Rho-dependent transcription termination process of bacteria.

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          Most cited references10

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          Regulator trafficking on bacterial transcription units in vivo.

          The trafficking patterns of the bacterial regulators of transcript elongation sigma(70), rho, NusA, and NusG on genes in vivo and the explanation for promoter-proximal peaks of RNA polymerase (RNAP) are unknown. Genome-wide, E. coli ChIP-chip revealed distinct association patterns of regulators as RNAP transcribes away from promoters (rho first, then NusA, then NusG). However, the interactions of elongating complexes with these regulators did not differ significantly among most transcription units. A modest variation of NusG signal among genes reflected increased NusG interaction as transcription progresses, rather than functional specialization of elongating complexes. Promoter-proximal RNAP peaks were offset from sigma(70) peaks in the direction of transcription and co-occurred with NusA and rho peaks, suggesting that the RNAP peaks reflected elongating, rather than initiating, complexes. However, inhibition of rho did not increase RNAP levels within genes downstream from the RNAP peaks, suggesting the peaks are caused by a mechanism other than rho-dependent attenuation.
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            An allosteric mechanism of Rho-dependent transcription termination.

            Rho is the essential RNA helicase that sets the borders between transcription units and adjusts transcriptional yield to translational needs in bacteria. Although Rho was the first termination factor to be discovered, the actual mechanism by which it reaches and disrupts the elongation complex (EC) is unknown. Here we show that the termination-committed Rho molecule associates with RNA polymerase (RNAP) throughout the transcription cycle; that is, it does not require the nascent transcript for initial binding. Moreover, the formation of the RNAP-Rho complex is crucial for termination. We show further that Rho-dependent termination is a two-step process that involves rapid EC inactivation (trap) and a relatively slow dissociation. Inactivation is the critical rate-limiting step that establishes the position of the termination site. The trap mechanism depends on the allosterically induced rearrangement of the RNAP catalytic centre by means of the evolutionarily conserved mobile trigger-loop domain, which is also required for EC dissociation. The key structural and functional similarities, which we found between Rho-dependent and intrinsic (Rho-independent) termination pathways, argue that the allosteric mechanism of termination is general and likely to be preserved for all cellular RNAPs throughout evolution.
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              Structure and folding of a designed knotted protein.

              A very small number of natural proteins have folded configurations in which the polypeptide backbone is knotted. Relatively little is known about the folding energy landscapes of such proteins, or how they have evolved. We explore those questions here by designing a unique knotted protein structure. Biophysical characterization and X-ray crystal structure determination show that the designed protein folds to the intended configuration, tying itself in a knot in the process, and that it folds reversibly. The protein folds to its native, knotted configuration approximately 20 times more slowly than a control protein, which was designed to have a similar tertiary structure but to be unknotted. Preliminary kinetic experiments suggest a complicated folding mechanism, providing opportunities for further characterization. The findings illustrate a situation where a protein is able to successfully traverse a complex folding energy landscape, though the amino acid sequence of the protein has not been subjected to evolutionary pressure for that ability. The success of the design strategy--connecting two monomers of an intertwined homodimer into a single protein chain--supports a model for evolution of knotted structures via gene duplication.
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                Author and article information

                Journal
                Bacteriophage
                Bacteriophage
                BACT
                Bacteriophage
                Landes Bioscience
                2159-7073
                2159-7081
                01 April 2013
                01 April 2013
                01 April 2013
                : 3
                : 2
                : e25657
                Affiliations
                [1 ]Laboratory of Transcription Biology; Center for DNA Fingerprinting and Diagnostics; Nampally, Hyderabad India
                [2 ]Crystallography and Molecular Biology Division; Saha Institute of Nuclear Physics; Bidhannagar, Kolkata India
                Author notes
                [†]

                Current affiliation: Department of Biochemistry; New York University School of Medicine; New York, NY USA

                [* ]Correspondence to: Ranjan Sen, Email: rsen@ 123456cdfd.org.in
                Article
                2013BACTERIOPHAGE0015R 25657
                10.4161/bact.25657
                3821671
                47fb8556-08e8-40d9-88ac-8021a4442a97
                Copyright © 2013 Landes Bioscience

                This is an open-access article licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported License. The article may be redistributed, reproduced, and reused for non-commercial purposes, provided the original source is properly cited.

                History
                : 07 June 2013
                : 04 July 2013
                : 08 July 2013
                Categories
                Article Addendum

                Microbiology & Virology
                bacteriophage,psu,rho,transcription termination
                Microbiology & Virology
                bacteriophage, psu, rho, transcription termination

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