Towards Understanding Helicase and Chaperone Activities in the RNA Degradosome

The E. coli RNA degradosome is a complex multi-enzyme machine which is central to the post-transcriptional regulation of the cell. Some of its functions include maturing and processing sRNA, rRNA, and tRNA, as well as degrading mRNA. The key components of the RNA degradosome include the endoribonuclease RNase E, the DEAD-box RNA helicase RhlB, the glycolytic enzyme enolase, and the phosphorolytic exoribonuclease PNPase. The degradosome has also been previously shown to associate with the RNA chaperone Hfq to form a small RNA guided machinery that targets defined transcripts. This thesis attempts to investigate several characteristics of the degradosome, including the importance of RhlB, the structure of a portion of the RNase E C-terminal domain that recruits enolase and helicase, and the association between Hfq, ChiX and the RNase E C-terminus. The thesis also explored structural details regarding the small domain of RNase E involved in both RNA binding and oligomerisation and its relationship to the RNA binding KH domains. Utilizing a point mutation in the DEAD box of RhlB, I have found increased RNA affinity to mutant RhlB, the potential structure of Hfq — an RNA chaperone — bound to the sRNA ChiX, and a structural correlation between the RNase E small domain and other KH domains. Preliminary models of Hfq and ChiX structure show a novel binding mode for class II sRNAs as the majority of ChiX associating with the distal face of Hfq. Bioinformatic studies reveal evolutionary roots between the KH domains and the RNase E small domain, supporting the hypothesis that the RNase E small domain may be involved in a novel mode of RNA binding and recognition.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2020

Author dc:creator

• Shen, Zi Ran

Advisor dc:contributor.advisor

• Luisi, Ben

• sRNA
• RNA degradosome
• RNase E
• PNPase
• KH domain
• ChiX
• Hfq
• RhlB

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/c61661e6-5025-46ca-bf6d-d9ccedd470af/download
• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

en

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.48562

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/301490