Collision Induced Unfolding Reveals Disease-Associated Stability Shifts in Mitochondrial tRNAs

24 August 2023, Version 1
This content is a preprint and has not undergone peer review at the time of posting.

Abstract

Ribonucleic acids (RNAs) remain challenging targets for structural biology, creating barriers to understanding their vast functions in cellular biology and fully realizing their applications in biotechnology. The inherent dynamism of RNAs creates numerous obstacles in capturing their biologically relevant higher-order structures (HOSs), and as a result, many RNA functions remain unknown. In this report, we describe the development of native ion mobility-mass spectrometry (IM-MS) and collision induced unfolding (CIU) for the structural characterization of a variety of RNAs. We evaluate the ability of these techniques to preserve native structural features into the gas-phase across a wide range of functional RNAs. Finally, we apply these tools to study the elusive mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS)-associated A3243G mutation. Our data demonstrate that our experimentally determined conditions preserve some solution-state memory of RNA via the increasing complexity of CIU fingerprints by RNA HOS and the retention of predicted magnesium binding events. Relating to the role of MELAS-inducing mutations in mitochondrial trans-fer RNA (mt-tRNAs), significant differences in collision cross-section (CCS) and stability are observed as a function of the A3243G mutation across a subset of the mitochondrial transfer RNA (tRNA) maturation pathway. We conclude by discussing the potential application of CIU for the development of RNA-based biotherapeutics and, more broadly, transcriptomic characterization.

Keywords

Ion Mobility
Native Mass Spectrometry
Nucleic Acid
Structural Biology

Supplementary materials

Title
Description
Actions
Title
Collision Induced Unfolding Reveals Disease-Associated Stability Shifts in Mitochondrial tRNAs
Description
RNA Sequence Information, Additional IM, MS, and CIU datasets, instrument settings, and RNA structure schematics
Actions

Comments

Comments are not moderated before they are posted, but they can be removed by the site moderators if they are found to be in contravention of our Commenting Policy [opens in a new tab] - please read this policy before you post. Comments should be used for scholarly discussion of the content in question. You can find more information about how to use the commenting feature here [opens in a new tab] .
This site is protected by reCAPTCHA and the Google Privacy Policy [opens in a new tab] and Terms of Service [opens in a new tab] apply.