RNA Structure Determination by Structural Probing and Mass Spectrometry: MS3D
Recent advances of detection strategies based on mass spectrometry (MS) have reawakened the interest in chemical methods for RNA structural elucidation by enabling experimental protocols that minimize their typical pitfalls. At the same time, the development of ever more sophisticated modeling techniques has helped close the resolution gap by providing atomic-level details that were previously beyond reach. Here, we describe the integration of MS-assisted structural probing with appropriate computational techniques, which has been termed MS3D, and illustrate its application to the elucidation of RNA substrates of biological significance. We address typical concerns faced by probing applications and possible solutions supported by the MS platform. We describe strategies for translating sparse spatial constraints afforded by footprinting and cross-linking reagents into testable all-atom structures. We also discuss future advances that would take further advantage of the synergy between experimental and computational approaches to increase the accuracy of chemical methods and to expand their scope to progressively larger and more complex targets.
KeywordsMouse Mammary Tumor Virus Bifunctional Reagent Monofunctional Adduct Crick Pairing Substrate Dynamic
This work was supported by National Institutes of Health Grant GM643208 and National Science Foundation Grant CHE-0439067.
- Aleksandrov ML et al (1984) Extraction of ions from solutions under atmospheric pressure: a method of mass spectrometric analysis of bioorganic compounds. Dokl Akad Nauk 277:379–383Google Scholar
- Cotter RJ (1997) Time-of-flight mass spectrometry. Instrumentation and applications in biological research. ACS, Washington, DCGoogle Scholar
- Hannis JC, Muddiman DC (1999) Characterization of a microdialysis approach to prepare polymerase chain reaction products for electrospray ionization mass spectrometry using on-line ultraviolet absorbance measurements and inductively coupled plasma atomic emission spectroscopy. Rapid Commun Mass Spectrom 13:323–330CrossRefGoogle Scholar
- Metz DH, Brown GL (1969) The investigation of nucleic acid secondary structure by means of chemical modification with a carbodiimide reagent. I. The reaction between N-cyclohexyl-N′-beta-(4-methylmorpholinium)ethylcarbodiimide and model nucleotides. Biochemistry 8(6):2312–2328PubMedCrossRefGoogle Scholar
- Muddiman DC et al (1996) Charge-state reduction with improved signal intensity of oligonucleotides in electrospray ionization mass spectrometry. J Am Soc Mass Spectrom 7:697–706Google Scholar
- Rozenski J (1999) Mongo Oligo Mass Calculator, v2.06Google Scholar
- Tanaka K et al (1987) Detection of high mass molecules by laser desorption time-of-flight mass spectrometry. In: Proceedings of the second Japan-China joint symposium on mass spectrometry. Bando Press, OsakaGoogle Scholar
- Urlaub H et al (1997) Identification and sequence analysis of contact sites between ribosomal proteins and rRNA in Escherichia coli 30S subunits by a new approach using matrix-assisted laser desorption/ionization-mass spectrometry combined with N-terminal microsequencing. J Biol Chem 272(23):14547–14555PubMedCrossRefGoogle Scholar