Interactions of in vitro selected fluorogenic peptide aptamers with calmodulin
Interactions of in vitro selected fluorogenic peptide aptamers with calmodulin
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DOI：
10.1007/s10529-016-2257-2
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发表时间：
2016-11
影响因子：
2.7
通讯作者：
Yasodha Manandhar;Wei Wang;Jin Inoue;N. Hayashi;T. Uzawa;Yutaka Ito;T. Aigaki;Yoshihiro Ito
中科院分区：
文献类型：
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作者：
Yasodha Manandhar;Wei Wang;Jin Inoue;N. Hayashi;T. Uzawa;Yutaka Ito;T. Aigaki;Yoshihiro Ito
目的通过使用先前选择的钙调素（CaM）适配体（包括非天然荧光氨基酸7-硝基-2,1,3-苯并二唑），在与选择过程相同的条件下，研究适配体使用的重要性。结果在n -末端添加5个氨基酸进行筛选，并对CaM结合的亲和力和选择性进行了测试。表面等离子体共振和荧光测量表明，其中一个适配体的额外氨基酸显著提高了与CaM的结合亲和力，表明在相同条件下适配体的使用与选择过程的重要性。在先前报道的序列中没有观察到如此剧烈的亲和力改善。核磁共振数据表明，主要结合位点位于CaM的交流端，附加残基增强了与CaM的相互作用。结论我们发现，加入用于核糖体展示的共同序列，使所选肽的亲和力与先前报道的肽一样强。
ObjectivesWe examined the importance of aptamer usage under the same condition as the selection process by employing the previously selected aptamers for calmodulin (CaM) which includes a non-natural fluorogenic amino acid, 7-nitro-2,1,3-benzoxadiazole.ResultsWe added five amino acids at theN-terminus which was employed for the selection and then we tested the affinity and selectivity for CaM binding. Surface plasmon resonance and fluorescence measurements showed that the additional amino acids for one of the aptamers drastically improved binding affinity to CaM, indicating the importance of aptamer use under the same conditions as the selection process. Such drastic improvement in affinity was not observed for the sequence which had been reported previously. Nuclear magnetic resonance data identified that the primary binding site is located in aC-terminal of CaM and the additional residues enhance interactions with CaM.ConclusionsWe found that the addition of the common sequence, which was employed for ribosome display, makes the affinity of a selected peptide as strong as the previously reported peptide.