title: Construction of a single chain variable fragment antibody (scFv) against tetrodotoxin (TTX) and its interaction with TTX.
abstract: Tetrodotoxin (TTX) is a small molecular weight neurotoxin that occludes voltage-gated sodium channels in nerve and muscle tissue, resulting in respiratory paralysis and death. A high affinity antibody that can neutralize the toxicity of TTX is still lacking, so it is very important to prepare an antibody for TTX therapy and detection. In the present study, a chemical method was used to prepare the tetrodotoxin complete antigen, and a small amount, repeatedly immunity way was carried to immunize 4 mice. The amplified genes encoding monoclonal antibodies against TTX were used to construct the phage display library. After six rounds of biopanning, an antibody named scFv-T53 was characterized from clones showing high affinity and specific to TTX, and its affinity constant was 1.1 × 10(6) L/mol. Three dimensional structure of the scFv-T53 was constructed by computer modeling, and TTX was docked to the scFv-T53 model to obtain the structure of the binding complex. Two predicted essential amino acids, K183 and I189, were mutated to verify the theoretical model. Both mutants lost binding activity significantly against TTX as predicted by the theoretical model. Hence, the above results will be useful for screening the high affinity anti-TTX scFv mutants.
authors.affiliation: Key Laboratory of Pathogenic Fungi and Mycotxins of Fujian Province, The Ministry of Education Key Laboratory of Biopesticide and Chemical Biology, School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China.
authors.forename: Rongzhi
authors.initials: R
authors.lastname: Wang
authors.affiliation: Bioengineering and Bioscience Department, Fuzhou University, Fuzhou 350108, China.
authors.forename: Ailing
authors.initials: A
authors.lastname: Huang
authors.affiliation: Key Laboratory of Pathogenic Fungi and Mycotxins of Fujian Province, The Ministry of Education Key Laboratory of Biopesticide and Chemical Biology, School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China.
authors.forename: Licai
authors.initials: L
authors.lastname: Liu
authors.affiliation: Key Laboratory of Pathogenic Fungi and Mycotxins of Fujian Province, The Ministry of Education Key Laboratory of Biopesticide and Chemical Biology, School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China.
authors.forename: Shuangshuang
authors.initials: S
authors.lastname: Xiang
authors.affiliation: Key Laboratory of Pathogenic Fungi and Mycotxins of Fujian Province, The Ministry of Education Key Laboratory of Biopesticide and Chemical Biology, School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China.
authors.forename: Xiufeng
authors.initials: X
authors.lastname: Li
authors.affiliation: Key Laboratory of Pathogenic Fungi and Mycotxins of Fujian Province, The Ministry of Education Key Laboratory of Biopesticide and Chemical Biology, School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China.
authors.forename: Sumei
authors.initials: S
authors.lastname: Ling
authors.affiliation: Key Laboratory of Pathogenic Fungi and Mycotxins of Fujian Province, The Ministry of Education Key Laboratory of Biopesticide and Chemical Biology, School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Wang
authors.affiliation: Bioengineering and Bioscience Department, Fuzhou University, Fuzhou 350108, China. Electronic address: lvtun@yahoo.com.
authors.forename: Tun
authors.initials: T
authors.lastname: Lu
authors.affiliation: Key Laboratory of Pathogenic Fungi and Mycotxins of Fujian Province, The Ministry of Education Key Laboratory of Biopesticide and Chemical Biology, School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou 350002, China. Electronic address: wshyyl@sina.com.
authors.forename: Shihua
authors.initials: S
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D057134:Antibodies, Neutralizing / Q000737:chemistry*; D000915:Antibody Affinity; D001483:Base Sequence; D001665:Binding Sites; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019151:Peptide Library; D017354:Point Mutation; D020539:Sequence Analysis, Protein; D057127:Single-Chain Antibodies / Q000737:chemistry*; D013779:Tetrodotoxin / Q000276:immunology*
chemical_list: D057134:Antibodies, Neutralizing; D019151:Peptide Library; D057127:Single-Chain Antibodies; D013779:Tetrodotoxin
keywords: Computer modeling; Mutation; Tetrodotoxin (TTX); scFv