STRUCTURE-FUNCTION RELATION OF THE NH2-TERMINAL DOMAIN OF THE SEMLIKI-FOREST-VIRUS CAPSID PROTEIN

The capsid (C) protein of alphaviruses consists of two protein domains : a serine protease at the COOH terminus and an NH2-terminal domain wh ich is thought to interact with RNA in the virus nucleocapsid (NC). Th e latter domain is very rich in positively charged amino acid residues , In this work, we have introduced large deletions into the correspond ing region of a full-length cDNA clone of Semliki Forest virus, expres sed the transcribed RNA in BHK-21 cells, and monitored the autoproteas e activity of C, the formation of intracellular NCs, and the release o f infectious virus. Our results show that if the gene region encoding the whole NH2-terminal domain is removed, the expressed C protein frag ment cannot assemble into NCs and virus particles but it is still able to function as an autoprotease, Thus, these results underline the gen eral importance of the NH2-terminal domain in the virus assembly proce ss and furthermore show that the serine protease domain can function i ndependently of the NH2 terminus. Surprisingly, analysis of additional C protein deletion variants showed that not all of the NH2-terminal d omain is required for virus assembly, but large deletions involving up to one-third of its positively charged residues are still compatible with NC and virus formation. The fact that so much flexibility is allo wed in the structure of the NH2-terminal domain of C suggests that mos t of this region is involved in nonspecific interactions with the enca psidated RNA, probably through its positively charged amino acid resid ues.