Title
Nucleic Acid-Dependent Structural Transition of the Intrinsically Disordered N-Terminal Appended Domain of Human Lysyl-tRNA Synthetase
Author
Soon Bin Kwon, Ji Eun Yu, Chan Park, Jiseop Lee, Baik L. Seong
Year
2018
Journal
International Journal of Molecular Sciences
Abstract
Eukaryotic lysyl-tRNA synthetases (LysRS) have an N-terminal appended tRNA-interaction domain (RID) that is absent in their prokaryotic counterparts. This domain is intrinsically disordered and lacks stable structures. The disorder-to-order transition is induced by tRNA binding and has implications on folding and subsequent assembly into multi-tRNA synthetase complexes. Here, we expressed and purified RID from human LysRS (hRID) in Escherichia coli and performed a detailed mutagenesis of the appended domain. hRID was co-purified with nucleic acids during Ni-affinity purification, and cumulative mutations on critical amino acid residues abolished RNA binding. Furthermore, we identified a structural ensemble between disordered and helical structures in non-RNA-binding mutants and an equilibrium shift for wild-type into the helical conformation upon RNA binding. Since mutations that disrupted RNA binding led to an increase in non-functional soluble aggregates, a stabilized RNA-mediated structural transition of the N-terminal appended domain may have implications on the functional organization of human LysRS and multi-tRNA synthetase complexes in vivo.
Full Article
Instrument
J-700
Keywords
Circular dichroism, Secondary structure, RNA structure, Biochemistry