Expression, crystallization and preliminary crystallographic study of the functional mutant (N60K) of nonstructural protein 9 fromHuman coronavirus HKU1

Abstract

Human coronavirus HKU1(HCoV-HKU1), which mainly causes acute self-limited respiratory-tract infections, belongs to group A of theBetacoronavirusgenus. Coronavirus genomes encode 16 nonstructural proteins (nsp1–16), which assemble into a large replication–transcription complex mediating virus propagation. Nonstructural protein 9, which binds to the single-stranded DNA/RNA, has been shown to be indispensible for viral replication. Interestingly, a functional mutant (N60K) of nsp9 was identified to compensate for a 6 nt insertion mutation of the 3′-untranslated region (UTR), which is critical for viral RNA synthesis. It has been proposed that the N60K mutation may cause certain conformational changes of nsp9 to rescue the defective insertion mutant. To further investigate the underlying structural mechanism, the N60K mutant of nsp9 from HCoV-HKU1 was successfully crystallized in this study. The crystals diffracted to 2.6 Å resolution and belonged to space groupP212121, with unit-cell parametersa= 31.9,b= 85.0,c= 95.0 Å. Two molecules were identified per asymmetric unit.