Expression，Purification，Crystallization of Acetylcholine Binding Proteins from Lymnaea stagnalis in Bac-to-Bac System

Abstract

Abstract:

It was to construct a recombinant bacmid that expresses acetylcholine binding protein from Lymnaea stagnalis（Ls-AChBP）and express the Ls-AChBP functional genes in Bac-to-Bac expression system to obtain the crystal. Synthesized Ls-AChBP cDNA was inserted into the correct position of the baculovirus expression vector pFastBac1. After the positive recombinant bacmid was screened, the recombinant bacmid was transfected into the insect cells to generate restructured baculovirus for high expression of the recombinant protein Ls-AChBP（rLs-AChBP）, which was captured by nickel-charged resin, and further purified by gel filtration chromatography. The rLs-AChBP pentamer was obtained and performed crystallization by robot screen. Recombinant Ls-AChBP expression in Bac-to-Bac system was successfully and we got Ls-AChBP crystal. Ls-AChBP is one of the AchBPs. The crystal structure of rLs-AchBP is an available template of description of nAChRs structure and function.

Key words: Ls-AChBP, Bac-to-Bac expression system, Recombinant expression, Purification, Crystallization

Lin Bo, Meng Hailing, Wu Yong, Bing Hui, Zhangsun Dongting, Luo Sulan. Expression，Purification，Crystallization of Acetylcholine Binding Proteins from Lymnaea stagnalis in Bac-to-Bac System[J]. Biotechnology Bulletin, 2014, 0(8): 126-131.

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