抗多环芳烃小鼠单链抗体的纯化与鉴定

Purification and characterization of mouse single-chain antibody against polycyclic aromatic hydrocarbons.

作者信息

Ustinov Valentin A, Averjanov Anton V, Glushkov Andrey N

机构信息

a Laboratory of Biotechnology , The Institute of Human Ecology of Siberian Branch of Russian Academy of Science , Kemerovo , Russia.

出版信息

J Immunoassay Immunochem. 2014;35(4):428-41. doi: 10.1080/15321819.2014.885449.

DOI:10.1080/15321819.2014.885449

PMID:24547788

Abstract

Polycyclic aromatic hydrocarbons (PAH) such as benzo[a]pyren mainly induce lung cancer in humans. We characterized the mouse single chain antibody against benzo[a]pyren (pSh). pSh was expressed and purified as cellulose binding domain fusion (pSh-CBD). The pSh-CBD bound five different PAH with high affinity. The 18 amino acid linker connected pSh-CBD heavy and light chains provided correct protein folding. The KDs for pSh-CBD and polycyclic aromatic hydrocarbons were similar to KDs for monoclonal antibody, approximately 10(-8). Separately heavy and light chains of pSh-CBD did not interact with benzo[a]pyren. Previously defined eleven pSh-CBD aa involved to benzo[a]pyren binding were confirmed by mutagenesis.

摘要

多环芳烃（PAH）如苯并[a]芘主要诱发人类肺癌。我们对针对苯并[a]芘的小鼠单链抗体（pSh）进行了表征。pSh作为纤维素结合域融合蛋白（pSh-CBD）进行表达和纯化。pSh-CBD与五种不同的多环芳烃具有高亲和力结合。连接pSh-CBD重链和轻链的18个氨基酸接头可实现正确的蛋白质折叠。pSh-CBD与多环芳烃的解离常数（KDs）与单克隆抗体的KDs相似，约为10^(-8)。pSh-CBD的重链和轻链单独存在时不与苯并[a]芘相互作用。通过诱变证实了先前确定的参与苯并[a]芘结合的11个pSh-CBD氨基酸。