Crystallization and preliminary X-ray crystallographic analysis of a nonstructural protein 15 mutant from Human coronavirus 229E.
Nonstructural protein 15 (nsp15), also called endoribonuclease, is a gene product of open reading frame 1b (ORF 1b) in coronaviruses. It is an important enzyme in the transcription/replication process involved in discontinuous negative-strand RNA synthesis. In this work, mutants of nsp15 from Human coronavirus 229E (HCoV-229E) were made based on structural analysis of the homologous nsp15s in Severe acute respiratory syndrome coronavirus (SARS-CoV) and Mouse hepatitis virus (MHV). The I26A/N52A mutant of nsp15 was overexpressed, purified and crystallized, and this mutant led to a trimeric form rather than hexamers or monomers. Crystals of trimeric nsp15 were obtained by the hanging-drop vapour-diffusion method using polyethylene glycol as a precipitant and diffracted to 2.5 Å resolution. The crystals belonged to space group C2221, with unit-cell parameters a = 85.9, b = 137.5, c = 423.1 Å, α = β = γ = 90°.
College of Life Sciences, Nankai University, Tianjin 300071, People's Republic of China.
Tong
T
Huo
College of Life Sciences, Nankai University, Tianjin 300071, People's Republic of China.
Xiang
X
Liu
D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D028941:Coronavirus 229E, Human / Q000737:chemistry*; D003460:Crystallization; D018360:Crystallography, X-Ray; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D008970:Molecular Weight; D050505:Mutant Proteins / Q000737:chemistry*; D014461:Ultracentrifugation; D017361:Viral Nonstructural Proteins / Q000737:chemistry*
D050505:Mutant Proteins; D017361:Viral Nonstructural Proteins
Human coronavirus 229E; coronaviruses; endoribonuclease; nonstructural protein 15; trimeric form