Recognition and cooperation between the ATP-dependent RNA helicase RhlB and ribonuclease RNase E |
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Authors: | Chandran Vidya Poljak Leonora Vanzo Nathalie F Leroy Anne Miguel Ricardo Núñez Fernandez-Recio Juan Parkinson James Burns Christopher Carpousis Agamemnon J Luisi Ben F |
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Affiliation: | Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK. |
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Abstract: | The Escherichia coli protein RhlB is an ATP-dependent motor that unfolds structured RNA for destruction by partner ribonucleases. In E. coli, and probably many other related gamma-proteobacteria, RhlB associates with the essential endoribonuclease RNase E as part of the multi-enzyme RNA degradosome assembly. The interaction with RNase E boosts RhlB's ATPase activity by an order of magnitude. Here, we examine the origins and implications of this effect. The location of the interaction sites on both RNase E and RhlB are refined and analysed using limited protease digestion, domain cross-linking and homology modelling. These data indicate that RhlB's carboxy-terminal RecA-like domain engages a segment of RNase E that is no greater than 64 residues. The interaction between RhlB and RNase E has two important consequences: first, the interaction itself stimulates the unwinding and ATPase activities of RhlB; second, RhlB gains proximity to two RNA-binding sites on RNase E, with which it cooperates to unwind RNA. Our homology model identifies a pattern of residues in RhlB that may be key for recognition of RNase E and which may communicate the activating effects. Our data also suggest that the association with RNase E may partially repress the RNA-binding activity of RhlB. This repression may in fact permit the interplay of the helicase and adjacent RNA binding segments as part of a process that steers substrates to either processing or destruction, depending on context, within the RNA degradosome assembly. |
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Keywords: | AR2, arginine-rich region 2 of RNase E (EC 3.1.26), corresponding to residues 798 to 819 DMS, dimethylsuberimidate ODA, optimal docking area PNPase, polynucleotide phosphorylase (EC 2.7.7.8) RBD, RNA-binding domain of RNase E, corresponding to residues 604 to 688 REP, repetitive extragenic palindrome RhlB, RNA helicase B (EC 3.6.1) RhlB-CTD, carboxy terminal domain of RhlB helicase, corresponding to residues 267 to 421 RhlB-NTD, amino terminal domain of RhlB helicase, corresponding to residues 1 to 266 RhlB-ΔCT, derivative of RhlB lacking the arginine-rich carboxy-terminal residues 398 to 421 RNase E (628-843), a segment of ribonuclease RNase E corresponding to residues 628 to 843 RNase E (498-1061), segment of RNase E containing residues 1-26 and 498 to 1061 RNase E (ΔRBD, ΔAR2), is a derivative of RNase E (498-1061) in which the RBD and AR2 have been deleted |
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