Elucidation of the stability and functional regions of the human coronavirus OC43 nucleocapsid protein.
Human coronavirus OC43 (HCoV-OC43) is one of the causes of the "common cold" in human during seasons of cold weather. The primary function of the HCoV-OC43 nucleocapsid protein (N protein) is to recognize viral genomic RNA, which leads to ribonucleocapsid formation. Here, we characterized the stability and identified the functional regions of the recombinant HCoV-OC43 N protein. Circular dichroism and fluorescence measurements revealed that the HCoV-OC43 N protein is more highly ordered and stabler than the SARS-CoV N protein previously studied. Surface plasmon resonance (SPR) experiments showed that the affinity of HCoV-OC43 N protein for RNA was approximately fivefold higher than that of N protein for DNA. Moreover, we found that the HCoV-OC43 N protein contains three RNA-binding regions in its N-terminal region (residues 1-173) and central-linker region (residues 174-232 and 233-300). The binding affinities of the truncated N proteins and RNA follow the order: residues 1-173-residues 233-300 > residues 174-232. SPR experiments demonstrated that the C-terminal region (residues 301-448) of HCoV-OC43 N protein lacks RNA-binding activity, while crosslinking and gel filtration analyses revealed that the C-terminal region is mainly involved in the oligomerization of the HCoV-OC43 N protein. This study may benefit the understanding of the mechanism of HCoV-OC43 nucleocapsid formation.
Huang
Chun-Yu
CY
Biotechnology Center, National Chung Hsing University, Taichung 402, Taiwan.
Hsu
Yen-Lan
YL
Chiang
Wan-Ling
WL
Hou
Ming-Hon
MH
D001665:Binding Sites; D002942:Circular Dichroism; D000086462:Coronavirus Nucleocapsid Proteins; D028962:Coronavirus OC43, Human / Q000737:chemistry*; D005976:Glutaral / Q000737:chemistry; D006801:Humans; D009696:Nucleic Acids / Q000378:metabolism; D019590:Nucleocapsid Proteins / Q000737:chemistry / Q000378:metabolism*; D055503:Protein Multimerization; D055550:Protein Stability; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D014508:Urea / Q000737:chemistry
D000086462:Coronavirus Nucleocapsid Proteins; D009696:Nucleic Acids; D019590:Nucleocapsid Proteins; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D014508:Urea; D005976:Glutaral
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