UCL Discovery
UCL home » Library Services » Electronic resources » UCL Discovery

Investigation of the Molecular Basis for Transcriptional Regulation of Tn916 and Macrolide Resistance in Bacillus subtilis

Mohamad Jamil, Norashirene Binti; (2020) Investigation of the Molecular Basis for Transcriptional Regulation of Tn916 and Macrolide Resistance in Bacillus subtilis. Doctoral thesis (Ph.D), UCL (University College London). Green open access

[thumbnail of NBMJ Thesis Corrected.pdf]
Preview
Text
NBMJ Thesis Corrected.pdf - Submitted Version

Download (27MB) | Preview

Abstract

Antibiotic resistance (AR) is one of the most serious threats to modern healthcare today. To understand how resistance spreads, we need to investigate the genetic basis of transferable AR. Conjugative transposons (CTns) have acquired the vast majority of resistance genes we currently know about which makes them one of the major vectors involved in their spread. This study aims to investigate how Tn916 and Tn916-like elements maintain their stability following insertion into a bacterial genome. We identified putative rho-independent terminators upstream of the conjugation genes of Tn2010, Tn5397, Tn6000, Tn6002, Tn6003, Tn6087 and Tn916 and hypothesised that their role is to prevent transcriptional readthrough into the conjugation genes upon integration into a new insertion site. To verify this experimentally, the terminator was cloned in between the tet(M) promoter and a gusA reporter in pHCMC05. We demonstrated the level of β-glucuronidase enzyme activity decreased, confirming termination activity. We have for the first time, identified and verified a group of conserved terminators in the conjugation region of the Tn916-like family of CTns. Further data supports our hypothesis that the terminator efficiency is modulated upon excision and circularisation of Tn916, which is the exact time when Tn916 would require expression of its conjugation genes. A fundamental understanding of the current antibiotic resistance mechanisms employed by bacteria is also essential to minimise the emergence of resistance and to devise effective resistance-control strategies. Another aim of this study is to investigate the molecular mechanism underlying macrolide resistance in Bacillus subtilis. Macrolide-resistant B. subtilis were generated as part of the project and analysis revealed a new genetic mutation to be responsible for the macrolide resistance phenotype. Comparative genome analysis revealed 21 bp and 54 bp duplication in the rplV of these mutants in comparison to the wild type strain. The rplV encodes the large ribosomal subunit protein, L22. Alteration in L22 has led to a predicted alteration in the C-terminal loop of the protein, predicted to change the shape of the exit tunnel within the ribosome. Ectopic expression of the rplV mutants containing the 21 bp and 54 bp duplication in B. subtilis BS34A confers resistance to macrolides. This is the first observation of macrolide resistance due to 54 bp duplication in the B. subtilis rplV gene.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Investigation of the Molecular Basis for Transcriptional Regulation of Tn916 and Macrolide Resistance in Bacillus subtilis
Event: UCL (University College London)
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Copyright © The Author 2020. Original content in this thesis is licensed under the terms of the Creative Commons Attribution 4.0 International (CC BY 4.0) Licence (https://creativecommons.org/licenses/by/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request.
UCL classification: UCL
UCL > Provost and Vice Provost Offices
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Eastman Dental Institute
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Eastman Dental Institute > Microbial Diseases
URI: https://discovery.ucl.ac.uk/id/eprint/10111989
Downloads since deposit
38Downloads
Download activity - last month
Download activity - last 12 months
Downloads by country - last 12 months

Archive Staff Only

View Item View Item