Surface plasmon resonance (SPR) biosensor is a promising technology for its various advantages including the real-time measurement of biomolecular interactions without labeling. A method of hybridizing RNAs on the surface of the streptavidin-coated (SA) sensor chip to study RNA-protein interactions was described in this paper. In our study, it has been shown that the nucleocapsid (N) protein of severe acute respiratory syndrome coronavirus (SARS-CoV) has a high binding affinity for the leader sequence of SARS-CoV genome. Effect of temperature on the RNA-DNA hybridization was also examined. This method can provide the affinity of interactions with high sensitivity. Therefore, it will be useful in screening binding candidates for a given RNA target motif with one chip.

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The 3'-one-third of the severe acute respiratory syndrome coronavirus (SARS-CoV) genome contains genes for four essential structural proteins and eight virus-specific genes. The expression of this genomic information of SARS-CoV involves synthesis of a nested set of subgenomic RNAs (sgRNAs). In this study, we showed that the translational levels of 10 SARS-CoV sgRNAs including the two low-abundance sgRNAs 2-1 and 3-1 varied considerably in translation reporter assays. We also demonstrated that the initiator AUG codon of sgRNA-8 was silent and the repressive control was most likely positioned in the upstream untranslated region (UTR) of sgRNA-8. The initiator AUG codons of most sgRNAs are in poor Kozak contexts and the translation of truncated proteins from downstream AUG codons by leaky scanning was common in our experimental settings. No significant correlation was found between complexity of 5'-UTR and the sequence context of AUG codon with the level of translation of SARS-CoV sgRNAs. These results will be helpful for further studies to reveal the biological functions and translation regulatory mechanisms of sgRNAs in the coronavirus life cycle and pathogenesis.

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