A novel method for characterizing the multi-functional C-terminal domain of the Hepadnavirus core protein

Abstract

The Hepadnavirus core protein is a viral structural protein with an N-terminal self-assembling domain and a C-terminal protamine-like arginine-rich domain (ARD). The ARD contains four clusters of arginine residues involved in RNA binding, genome DNA synthesis, and nuclear localization. Characterization of the multi-functions of ARD has been impeded due to the insoluble nature of the core protein expressed in vitro. A GST (glutathione-S-transferase) and ARD fusion protein, GST–ARD, was expressed and purified in this study. Gel mobility shift assays using purified GST–ARD fusion proteins demonstrated that, similar to protamine, the ARD domain of the core protein bound to oligonucleotides without sequence preference. In vitro affinity chromatography binding assays showed further that the ARD bound to tested random plasmid DNA in a sequence-independent manner. The GST–ARD fusion protein-based approach can be employed further to study other biochemical properties of the core protein.