Low stability of nucleocapsid protein in SARS virus.
The nucleocapsid protein (N protein) is one of the major virion structural proteins of a newly identified coronavirus, which has been confirmed as the causative agent of severe acute respiratory syndrome (SARS). The major function of N protein is to assemble the RNA of coronavirus. In the present study, the gene encoding the N protein was cloned and the protein was expressed, purified, and refolded as shown by (1)H NMR measurement. The maximal Trp emission wavelength occurs near 331 nm, suggesting substantial burial of Trp residues. Circular dichroism measurements indicate that N protein contains little alpha-helical structure. Acid titration shows that N protein begins to unfold near pH 5 and is fully denatured near pH 2.7, and the acid unfolding process is reversible. The physical and chemical properties of N protein indicate that its stability is low. N protein is denatured reversibly at pH 7.4 either by urea (with C(m) of 2.77 M and m value of 2.74 kcal mol(-1) M(-1)) or GdmCl (with C(m) of 1.46 M and m value of 4.50 kcal mol(-1) M(-1)). In the heat-induced denaturation in phosphate-buffered saline buffer, N-protein starts to unfold at 35 degrees C and is completely denatured at 55 degrees C, where SARS virus was also reported to be inactivated. We propose that the low stability of N protein may be critical for the stability and function of SARS virus.
Department of Biological Sciences and Biotechnology, MOE Laboratory of Protein Science, Tsinghua University, Beijing 100084, People's Republic of China.
Yulong
Y
Wang
Xiaoyu
X
Wu
Yihua
Y
Wang
Bing
B
Li
Hao
H
Zhou
Guiyong
G
Yuan
Yan
Y
Fu
Yongzhang
Y
Luo
D002414:Cations, Monovalent; D019791:Guanidine / Q000737:chemistry; D006358:Hot Temperature; D006801:Humans; D006863:Hydrogen-Ion Concentration; D019590:Nucleocapsid Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D011489:Protein Denaturation; D017510:Protein Folding; D045473:Severe acute respiratory syndrome-related coronavirus / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D045169:Severe Acute Respiratory Syndrome / Q000821:virology; D012965:Sodium Chloride / Q000737:chemistry; D013816:Thermodynamics; D014508:Urea / Q000737:chemistry; D038301:Virus Inactivation
D002414:Cations, Monovalent; D019590:Nucleocapsid Proteins; D012965:Sodium Chloride; D014508:Urea; D019791:Guanidine