Estimation of interaction between oriented immobilized green fluorescent protein and its antibody by high performance affinity chromatography and molecular docking.
Although green fluorescence protein (GFP) and its antibody are widely used to track a protein or a cell in life sciences, the binding behavior between them remains unclear. In this work, diazo coupling method that synthesized a new stationary GFP was oriented immobilized on the surface of macro-porous silica gel by a phase. The stationary phase was utilized to confirm the validation of injection amount-dependent analysis in exploring protein-protein interaction that use GFP antibody as a probe. GFP antibody was proved to have one type of binding site on immobilized GFP. The number of binding site and association constant were calculated to be (6.41 ± 0.76) × 10(-10) M and (1.39 ± 0.12) × 10(9) M(-1). Further analysis by molecular docking showed that the binding of GFP to its antibody is mainly driven by hydrogen bonds and salt bridges. These results indicated that injection amount-dependent analysis is capable of exploring the protein-protein interactions with the advantages of ligand and time saving. It is a valuable methodology for the ligands, which are expensive or difficult to obtain.
College of Life Sciences, Northwest University, Xi'an, 710069, China.
Qian
Q
Li
College of Life Sciences, Northwest University, Xi'an, 710069, China.
Jing
J
Wang
College of Life Sciences, Northwest University, Xi'an, 710069, China.
Lingjian
L
Yang
College of Life Sciences, Northwest University, Xi'an, 710069, China.
Xiaokang
X
Gao
College of Life Sciences, Northwest University, Xi'an, 710069, China.
Hongwei
H
Chen
College of Life Sciences, Northwest University, Xi'an, 710069, China.
Xinfeng
X
Zhao
College of Life Sciences, Northwest University, Xi'an, 710069, China.
Liujiao
L
Bian
College of Life Sciences, Northwest University, Xi'an, 710069, China.
Xiaohui
X
Zheng
D000906:Antibodies / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods; D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000378:metabolism; D062105:Molecular Docking Simulation / Q000379:methods; D011485:Protein Binding
D000906:Antibodies; D049452:Green Fluorescent Proteins
affinity chromatography; antigen-antibody interaction; diazotization; molecular docking; oriented immobilization; protein-protein interaction