Analysis of the relationship between end-to-end distance and activity of single-chain antibody against colorectal carcinoma.
We investigated the relationship of End-to-end distance between VH and VL with different peptide linkers and the activity of single-chain antibodies by computer-aided simulation. First, we developed (G4S)n (where n = 1-9) as the linker to connect VH and VL, and estimated the 3D structure of single-chain Fv antibody (scFv) by homologous modeling. After molecular models were evaluated and optimized, the coordinate system of every protein was built and unified into one coordinate system, and End-to-end distances calculated using 3D space coordinates. After expression and purification of scFv-n with (G4S)n as n = 1, 3, 5, 7 or 9, the immunoreactivity of purified ND-1 scFv-n was determined by ELISA. A multi-factorial relationship model was employed to analyze the structural factors affecting scFv: rn=ABn-ABO2+CDn-CDO2+BCn-BCst2. The relationship between immunoreactivity and r-values revealed that fusion protein structure approached the desired state when the r-value = 3. The immunoreactivity declined as the r-value increased, but when the r-value exceeded a certain threshold, it stabilized. We used a linear relationship to analyze structural factors affecting scFv immunoreactivity.
Faculty of Biomedical Engineering of Zhengzhou University, Zhengzhou, 450001, Henan Province, People's Republic of China.
Jianhua
J
Zhang
Shanhong
S
Liu
Zhigang
Z
Shang
Li
L
Shi
Jun
J
Yun
D000595:Amino Acid Sequence; D001483:Base Sequence; D015153:Blotting, Western; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000276:immunology*; D003198:Computer Simulation; D017931:DNA Primers; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D008969:Molecular Sequence Data; D057127:Single-Chain Antibodies / Q000737:chemistry / Q000276:immunology*
D017931:DNA Primers; D057127:Single-Chain Antibodies