Poster abstracts
Poster number 57 submitted by Jeff Levengood
Utilization of high-pressure NMR for the study of the folded and disordered domains of hnRNP A1
Jeffrey D. Levengood (Department of Chemistry, Case Western Reserve University), Julien Roche (Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University), Blanton S. Tolbert (Department of Chemistry, Case Western Reserve University)
Abstract:
The hnRNP A1 protein is involved in numerous processes of nascent RNA transcripts. These roles include, but are not limited to, translational control, splicing regulation, and mRNA stabilization. The protein performs its functions by binding RNA at a high affinity site before recruiting other proteins to aid in its RNA processing activities.
hnRNP A1 is composed of three domains, two structurally identical RRMs that form the N-terminal protein UP1, and a disordered, glycine rich C-terminal domain (G-CTD). The UP1 domain is responsible for binding the nucleic acid substrates while the G-CTD assembles the multi-protein complexes the protein utilizes to carry out its various functions. It is also a low-complexity domain (LCD) capable of mediating liquid-liquid phase separation.
High-pressure NMR spectroscopy is a powerful technique for examining the biophysical nature of proteins. Application of pressure to a protein increases solvent density and pushes the protein towards a lower molar volume, causing folded proteins to unfold. For disordered domains, pressure has been shown to have just minor effects on NMR parameters.
The utilization of high-pressure NMR spectroscopy for the study of full length hnRNP A1 revealed unique properties for both the structured and disordered domains. It revealed the LCD, at standard conditions, adopts a disordered, but compact conformation that is stabilized by polar and electrostatic interactions. The application of pressure results in a disruption of the intra-protein interactions and the compact nature of the domain.
While pressure normally unfolds proteins, the structured UP1 domain failed to unfold at high pressure. To study this phenomenon, UP1 variants were examined to determine what properties of the protein are responsible for its stability at high pressure. Results have revealed that communication between the two RRM domains is vital for maintenance of the protein’s stability during the application of high-pressure.
References:
Levengood JD, Peterson J, Tolbert BS, Roche J. Thermodynamic stability of hnRNP A1 low complexity domain revealed by high-pressure NMR. Proteins. 2021 Jul;89(7):781-791.
Keywords: RRM, IDP, high-pressure NMR