Pirooznia Nazanin, Hasannia Sadegh, Taghdir Majid, Rahbarizadeh Fatemeh, Eskandani Morteza
Department of Biology, Faculty of Science, University of Guilan, Rasht, Iran.
J Biomed Biotechnol. 2011;2011:578128. doi: 10.1155/2011/578128. Epub 2011 Aug 22.
Adaptive cell immunotherapy with the use of chimeric receptors leads to the best and most specific response against tumors. Chimeric receptors consist of a signaling fragment, extracellular spacer, costimulating domain, and an antibody. Antibodies cause immunogenicity; therefore, VHH is a good replacement for ScFv in chimeric receptors. Since peptide sequences have an influence on chimeric receptors, the effect of peptide domains on each other's conformation were investigated. CD3Zeta, CD28, VHH and CD8α, and FcgIIα are used as signaling moieties, costimulating domain, antibody, and spacers, respectively. To investigate the influence of the ligation of spacers on the conformational structure of VHH, models of VHH were constructed. Molecular dynamics simulation was run to study the influence of the presence of spacers on the conformational changes in the binding sites of VHH. Root mean square deviation and root mean square fluctuation of critical segments in the binding site showed no noticeable differences with those in the native VHH. Results from molecular docking revealed that the presence of spacer FcgIIα causes an increasing effect on VHH with MUC1 interaction. Each of the constructs was transformed into the Jurkat E6.1. Expression analysis and evaluation of their functions were examined. The results showed good expression and function.
使用嵌合受体的适应性细胞免疫疗法可产生针对肿瘤的最佳且最具特异性的反应。嵌合受体由信号片段、细胞外间隔区、共刺激结构域和抗体组成。抗体具有免疫原性；因此，VHH是嵌合受体中ScFv的良好替代物。由于肽序列会影响嵌合受体，因此研究了肽结构域对彼此构象的影响。分别使用CD3Zeta、CD28、VHH和CD8α以及FcgIIα作为信号部分、共刺激结构域、抗体和间隔区。为了研究间隔区连接对VHH构象结构的影响，构建了VHH模型。进行分子动力学模拟以研究间隔区的存在对VHH结合位点构象变化的影响。结合位点关键片段的均方根偏差和均方根波动与天然VHH中的相比没有明显差异。分子对接结果表明，间隔区FcgIIα的存在对VHH与MUC1的相互作用有增强作用。将每个构建体转化到Jurkat E6.1中。检查了它们的表达分析和功能评估。结果显示表达和功能良好。
