Complementarity determining regions of an anti-prion protein scFv fragment orchestrate conformation specificity and antiprion activity
Complementarity determining regions of an anti-prion protein scFv fragment orchestrate conformation specificity and antiprion activity
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DOI：
10.1016/j.molimm.2008.07.023
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发表时间：
2009-02-01
影响因子：
3.6
通讯作者：
Korth, Carsten
中科院分区：
文献类型：
--
作者：
Mueller-Schiffmann, Andreas;Petsch, Benjamin;Korth, Carsten
Prion蛋白PrP以几种稳定的构象存在，其中一种构象PrPSc与遗传性神经退行性疾病有关。通过高亲和力配体靶向PrP已被证明是预防外周PrP感染的有效方法。在这里，我们已经在大肠杆菌中产生了单抗可变区(ScFv)的细菌表达单链片段，最初是针对识别PrPc和PrPSc的纯化PrPSc而提出的。该单链抗体与重组PrP的解离常数(K-D)为2 nM，并在4 nM处清除了SCN2A细胞中的PrP。与重链互补决定区3(CDR3H)相对应的肽选择性地与PrPSc结合，但失去了抗朊病毒活性。然而，一种模拟CDR3H侧链拓扑结构的改进多肽的合成和应用，在显示出更强的蛋白酶抗性的同时，仍然结合了PrPSc和恢复了抗蛋白原活性。我们得出的结论是：(1)scFvW226是迄今为止生物测定证实的具有抗病毒活性的最小多肽，(2)不同CDR的参与可以调节不同的构象特异性和生物活性。(C)2008爱思唯尔有限公司。保留所有权利。
The prion protein, PrP, exists in several stable conformations, with the presence of one conformation, PrPSc, associated with transmissible neurodegenerative diseases. Targeting PrP by high-affinity ligands has been proven to be an effective way of preventing peripheral prion infections. Here, we have generated bacterially expressed single chain fragments of the variable domains (scFv) of a monoclonal antibody in Escherichia coli, originally raised against purified PrPSc that recognizes both PrPc and PrPSc. This scFv fragment had a dissociation constant (K-D) with recombinant PrP of 2 nM and cleared prions in ScN2a cells at 4 nM, as demonstrated by a mouse prion bioassay. A peptide corresponding to the complementarity determining region 3 of the heavy chain (CDR3H) selectively bound PrPSc but had lost antiprion activity. However, synthesis and application of an improved peptide mimicking side chain topology of CDR3H while exhibiting increased protease resistance, a retro-inverso D-peptide of CDR3H, still bound PrPSc and reinstated antiprion activity. We conclude that (1) scFvW226 is so far the smallest polypeptide with bioassay confirmed antiprion activity, and (2) differential conformation specificity and bioactivity can be regulated by orchestrating the participation of different CDRs. (C) 2008 Elsevier Ltd. All rights reserved.