Hepatitis B Virus Core Protein Phosphorylation Sites Affect Capsid Stability and Transient Exposure of the C-terminal Domain [Protein Structure and Folding]
September 24th, 2015 by Selzer, L., Kant, R., Wang, J. C.-Y., Bothner, B., Zlotnick, A.
Hepatitis B Virus core protein has 183 amino acids divided into an assembly domain and an arginine-rich C-terminal domain (CTD) that regulates essential functions including genome packaging, reverse transcription, and intracellular trafficking. Here, we investigated the CTD in empty HBV T=4 capsids. We examined wild-type core protein (Cp183-WT) and a mutant core protein (Cp183-EEE), in which three CTD serines are replaced with glutamate to mimic phosphorylated protein. We found that Cp183-WT capsids were less stable than Cp183-EEE capsids. When we tested CTD sensitivity to trypsin, we detected two different populations of CTDs differentiated by their rate of trypsin cleavage. Interestingly, CTDs from Cp183-EEE capsids exhibited a much slower rate of proteolytic cleavage compared to CTDs of Cp183-WT capsids. Cryo-EM studies of trypsin-digested capsids show that CTDs at fivefold symmetry vertices are most protected. We hypothesize that electrostatic interactions between glutamates and arginines in Cp183-EEE, particularly at fivefolds, increase capsid stability and reduce CTD exposure. Our studies show that quasi-equivalent CTDs exhibit different rates of exposure and thus might perform distinct functions during the HBV lifecycle. Our results demonstrate a structural role for CTD phosphorylation and indicate crosstalk between CTDs within a capsid particle.