NMR characterisation of a novel neamine antibiotic 12 and its interaction with a conserved 27mer RNA motif of 16S rRNA

Abstract

In recent times, the growing challenge of antibiotic resistance has prompted intense efforts to elucidate the mechanism of action of antibiotics at the molecular level, using techniques such as NMR spectroscopy. Blocking protein synthesis is an effective way of combating bacterial infection and many antibiotics function in this manner. Interest in the involvement of RNA in protein biosynthesis has increased following extensive studies on the binding of antibiotic drugs to specific target sites on ribosomal RNA. A systematic NMR study of a novel antibiotic derived from neamine and its interaction with a conserved and highly stable 27mer RNA motif of the A-site 16S rRNA was carried out. The antibiotic showed well resolved and dispersed resonances including exchange retarded amide protons suggesting a stable, folded conformation of the drug. The NMR results obtained provide a structural rationale to modify selected groups on the neamine 12 antibiotic to enhance the affinity for the RNA. The empirical Gibbs energy (DeltaG=-10.70 kcal/mol) and secondary structure were predicted for the 27mer RNA by the Vienna RNAfold software exhibiting a good thermodynamic stability. Similarly, the 27mer RNA exhibited well resolved and stable exchangeable imino and amino resonances in the lowfield region of the 1H-NMR spectrum. The NMR spectra of the 27mer RNA-neamine 12 complex showed small but detectable changes in chemical shift and linewidth, indicating weak interaction between the neamine 12 and the 27mer RNA. These changes can be qualitatively interpreted as changes in the local conformation of the 27mer RNA and the neamine 12, arising from the formation of their complex. The NMR results obtained have laid a solid foundation to determine the three dimensional solution state structures of neamine 12 after necessary modification and the complex with the RNA to elucidate the mechanism of action of the antibiotic.

Date of Award	31 Dec 2013
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Vasudevan Ramesh (Supervisor) & Raymond O'Keefe (Supervisor)