The nucleocapsid protein of SARS coronavirus has a high binding affinity to the human cellular heterogeneous nuclear ribonucleoprotein A1

The nucleocapsid (N) protein of SARS coronavirus (SARS_CoV) is a major structural component of virions, which appears to be a multifunctional protein involved in viral RNA replication and translation. Heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) is related to the pre-mRNA splicing in the nu...

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Bibliographic Details
Published in:	FEBS letters 2005-05, Vol.579 (12), p.2623-2628
Main Authors:	Luo, Haibin, Chen, Qing, Chen, Jing, Chen, Kaixian, Shen, Xu, Jiang, Hualiang
Format:	Article
Language:	eng
Subjects:	Clone Cells Electrophoresis, Polyacrylamide Gel Glutathione Transferase - metabolism Glycine - chemistry GST pull-down Heterogeneous Nuclear Ribonucleoprotein A1 Heterogeneous-Nuclear Ribonucleoprotein Group A-B - chemistry Heterogeneous-Nuclear Ribonucleoprotein Group A-B - metabolism hnRNP A1 Humans IPTG isopropyl-β-d-galactoside Kinetics MHV mouse hepatitis virus Murine hepatitis virus N protein Nucleocapsid protein Nucleocapsid Proteins - chemistry Nucleocapsid Proteins - genetics Nucleocapsid Proteins - isolation & purification Nucleocapsid Proteins - metabolism Peptide Fragments - metabolism Protein Structure, Tertiary Recombinant Proteins - metabolism RNA, Viral - biosynthesis Saccharomyces cerevisiae - growth & development Saccharomyces cerevisiae - metabolism SARS coronavirus SARS_CoV SARS_N Severe Acute Respiratory Syndrome - metabolism severe acute respiratory syndrome coronavirus Short Communication Short Communications SPR Surface Plasmon Resonance the nucleocapsid protein of SARS coronavirus Transformation, Genetic Two-Hybrid System Techniques Yeast two-hybrid
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Summary:	The nucleocapsid (N) protein of SARS coronavirus (SARS_CoV) is a major structural component of virions, which appears to be a multifunctional protein involved in viral RNA replication and translation. Heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) is related to the pre-mRNA splicing in the nucleus and translation regulation in the cytoplasm. In this report, based on the relevant biophysical and biochemical assays, the nucleocapsid protein of SARS_CoV (SARS_N) was discovered to exhibit high binding affinity to human hnRNP A1. GST pull-down results clearly demonstrated that SARS_N protein could directly and specifically bind to human hnRNP A1 in vitro. Yeast two-hybrid assays further indicated in vivo that such binding relates to the fragment (aa 161–210) of SARS_N and the Gly-rich domain (aa 203–320) of hnRNP A1. Moreover, kinetic analyses by surface plasmon resonance (SPR) technology revealed that SARS_N protein has a specific binding affinity against human hnRNP A1 with K D at 0.35 ± 0.02 μM ( k on = 5.83 ± 0.42 × 10 3 M −1 s −1 and k off = 2.06 ± 0.12 × 10 −3 s −1). It is suggested that both SARS_N and hnRNP A1 proteins are possibly within the SARS_CoV replication/transcription complex and SARS_N/human hnRNP A1 interaction might function in the regulation of SARS_CoV RNA synthesis. In addition, the determined results showed that SARS_N protein has only one binding domain for interacting with human hnRNP A1, which is different from the mouse hepatitis virus (MHV) binding case where the nucleocapsid protein of MHV (MHV_N) was found to have two binding domains involved in the MHV_N/hnRNP A1 interaction, thereby suggesting that SARS_N protein might carry out a different binding mode to bind to human hnRNP A1 for its further function performance in comparison with MHV_N.
ISSN:	0014-5793 1873-3468