Institute of Immunology, Zhongshan School of Medicine, Key Laboratory of Tropical Disease Control Research of Ministry of Education, Sun Yat-sen University, Guangzhou, China.
BMC Immunol. 2012 Sep 10;13:50. doi: 10.1186/1471-2172-13-50.
Severe acute respiratory syndrome (SARS) is an emerging infectious disease caused by the novel coronavirus SARS-CoV. The T cell epitopes of the SARS CoV spike protein are well known, but no systematic evaluation of the functional and structural roles of each residue has been reported for these antigenic epitopes. Analysis of the functional importance of side-chains by mutational study may exaggerate the effect by imposing a structural disturbance or an unusual steric, electrostatic or hydrophobic interaction.
We demonstrated that N50 could induce significant IFN-gamma response from SARS-CoV S DNA immunized mice splenocytes by the means of ELISA, ELISPOT and FACS. Moreover, S366-374 was predicted to be an optimal epitope by bioinformatics tools: ANN, SMM, ARB and BIMAS, and confirmed by IFN-gamma response induced by a series of S358-374-derived peptides. Furthermore, each of S366-374 was replaced by alanine (A), lysine (K) or aspartic acid (D), respectively. ANN was used to estimate the binding affinity of single S366-374 mutants to H-2 Kd. Y367 and L374 were predicated to possess the most important role in peptide binding. Additionally, these one residue mutated peptides were synthesized, and IFN-gamma production induced by G368, V369, A371, T372 and K373 mutated S366-374 were decreased obviously.
We demonstrated that S366-374 is an optimal H-2 Kd CTL epitope in the SARS CoV S protein. Moreover, Y367, S370, and L374 are anchors in the epitope, while C366, G368, V369, A371, T372, and K373 may directly interact with TCR on the surface of CD8-T cells.
严重急性呼吸系统综合症（SARS）是一种新型传染病，由新型冠状病毒 SARS-CoV 引起。SARS-CoV 刺突蛋白的 T 细胞表位众所周知，但尚未有报道对这些抗原表位的每个残基的功能和结构作用进行系统评估。通过突变研究分析侧链的功能重要性可能会通过施加结构干扰或不寻常的空间位阻、静电或疏水性相互作用而夸大效果。
我们通过 ELISA、ELISPOT 和 FACS 证明，N50 可以诱导 SARS-CoV S DNA 免疫的小鼠脾细胞产生显著的 IFN-γ反应。此外，生物信息学工具（ANN、SMM、ARB 和 BIMAS）预测 S366-374 是一个最佳的表位，并通过一系列 S358-374 衍生肽诱导 IFN-γ反应得到证实。此外，S366-374 中的每个残基分别被丙氨酸（A）、赖氨酸（K）或天冬氨酸（D）取代。ANN 用于估计单个 S366-374 突变体与 H-2 Kd 的结合亲和力。Y367 和 L374 被预测在肽结合中具有最重要的作用。此外，这些一个残基突变的肽被合成，并且 G368、V369、A371、T372 和 K373 突变的 S366-374 诱导的 IFN-γ产生明显减少。
我们证明 S366-374 是 SARS-CoV S 蛋白中最佳的 H-2 Kd CTL 表位。此外，Y367、S370 和 L374 是表位的锚点，而 C366、G368、V369、A371、T372 和 K373 可能直接与 CD8-T 细胞表面的 TCR 相互作用。
