Analysis of the Relationship Between Molecular Conformation of Single-Chain Variable Fragment and In Vitro Biological Activities by Modeling

摘要

Objective: To analyze the influence of different number of connecting peptides on antibody expression activity and obtain the number of the connecting peptides with which the expression activity is the most similarity to the wild-type. Methods: This study obtained single-chain variable fragment (ScFv) ND-1ScFv-n (n = 0~9) by constructing its prokaryotic expression vector. The similarity between the wild-type antibody and ScFv was obtained by space spherical shell layered method based on spherical coordinates, by which ND-1ScFv-n with the most similarity to the molecular conformation and biological activity of wild-type antibody was determined. The biological activity of ND-1ScFv-n was tested through ELISA and Western-blot. Results: When n = 3, the conformational similarity of the constructed ScFv and wild-type antibody was the highest and their biological activity was the closest. The regression equation of the relationship between the similarity (dependent variable y) and the terminal distance ratio of VH in ND-1ScFv-n and in the wild type, AB (ND-1ScFv-n)/AB (wild type) (independent variable x1) and the terminal distance ratio of VL in ND-1ScFv-n and in the wild type, CD (ND-1ScFv-n)/CD (wild type) (independent variable x2) was y = 0.8459+0.0088x1 -0.0098x2. Conclusion: When the number of connecting peptides was three, the conformation and biological activity of ScFv was the closest to those of wild-type antibody.