Abstract
Multi-domain proteins play critical roles in fine-tuning essential processes in cellular signaling and gene regulation. Typically, multiple globular domains that are connected by flexible linkers undergo dynamic rearrangements upon binding to protein, DNA or RNA ligands. RNA binding proteins (RBPs) represent an important class of multi-domain proteins, which regulate gene expression by recognizing linear or structured RNA sequence motifs. Here, we employ segmental perdeuteration of the three RNA recognition motif (RRM) domains in the RBP TIA-1 using Sortase A mediated protein ligation. We show that domain-selective perdeuteration combined with contrast-matched small-angle neutron scattering (SANS), SAXS and computational modeling provides valuable information to precisely define relative domain arrangements. The approach is generally applicable to study conformational arrangements of individual domains in multi-domain proteins and changes induced by ligand binding.
Highlights
Multi-domain proteins play critical roles in fine-tuning essential processes in cellular signaling and gene regulation
The domain arrangements and changes induced by ligand binding play important roles for the molecular function of these
A current model for the molecular functions of TIA-1 suggests that RRM2 and RRM3 bind to pre-mRNA, whereas RRM1 and the Q-rich domain interact with the U1C protein; a component of the spliceosomal U1 snRNP complex (Figure 1b).[13]
Summary
Multi-domain proteins play critical roles in fine-tuning essential processes in cellular signaling and gene regulation. The combination of solution NMR with small angle scattering is advantageous as NMR provides information on binding interfaces, domain conformations and dynamics, while small angle scattering experiments yield information about overall shapes.[4] Of special interest is the use of contrast matching[5] in SANS, where the location of individual subunits in subunitselectively perdeuterated complexes can be determined.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
