Systems NMR: single-sample quantification of RNA, proteins and metabolites for biomolecular network analysis

Nature Methods
Yaroslav NikolaevFrédéric H-T Allain

Abstract

Cellular behavior is controlled by the interplay of diverse biomolecules. Most experimental methods, however, can only monitor a single molecule class or reaction type at a time. We developed an in vitro nuclear magnetic resonance spectroscopy (NMR) approach, which permitted dynamic quantification of an entire 'heterotypic' network-simultaneously monitoring three distinct molecule classes (metabolites, proteins and RNA) and all elementary reaction types (bimolecular interactions, catalysis, unimolecular changes). Focusing on an eight-reaction co-transcriptional RNA folding network, in a single sample we recorded over 35 time points with over 170 observables each, and accurately determined five core reaction constants in multiplex. This reconstruction revealed unexpected cross-talk between the different reactions. We further observed dynamic phase-separation in a system of five distinct RNA-binding domains in the course of the RNA transcription reaction. Our Systems NMR approach provides a deeper understanding of biological network dynamics by combining the dynamic resolution of biochemical assays and the multiplexing ability of 'omics'.

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Citations

Jul 12, 2020·Journal of Biomolecular NMR·Andras BoeszoermenyiGerhard Wagner
Oct 28, 2020·Small·Md Sohel RanaKun Qian
Nov 4, 2020·Open Biology·Gitanjali AsampilleHanudatta S Atreya
Aug 10, 2021·Current Opinion in Structural Biology·Leonidas EmmanouilidisFrédéric H-T Allain
Nov 13, 2020·Analytical Chemistry·Arthur S EdisonSicong Zhang
Nov 16, 2021·The EMBO Journal·Andrew M JobbinsIan C Eperon

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Methods Mentioned

BETA
NMR
RNA folding
transcription-NMR
chemical shift

Software Mentioned

RNAfold
BioNetGen
Systems NMR
Affinimeter
rbnmr
MATLAB
MicroCal Origin
TopSpin
lorentzfit
Python

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