1. Introduction to surface remodeling antibodies

Surface remodeling antibodies refer to humanization of amino acid residues on the surface of heterologous antibodies. The principle of this method is to replace only the regions that are significantly different from the SAR of human antibodies, and select amino acid replacements similar to the surface residues of human antibodies on the basis of maintaining antibody activity and reducing heterology; in addition, the replaced segments should not be too many, and residues that affect the size, charge, hydrophobicity of the side chain, or may form hydrogen bonds and thus affect the conformation of the antibody complementary determining region (CDR) should not be replaced as much as possible.

2. Surface remodeling antibody method

By comparing the antibody variable region sequence with the mouse and human antibody variable region sequence library, the differences and abnormal residues are obtained. Further, based on the three-dimensional modeling and analysis of the intramolecular and intermolecular hydrogen bond sites and surface accessibility of the antibody variable region, the core residues involved in hydrogen bond formation and hydrophobicity are removed to complete the humanization of the antibody variable region on the principle of ensuring the integrity and affinity of the antibody structure.

3. Surface remodeling antibody preparation process (technical difficulties)

Analysis of antibody variable region sequence and humanized design

Use ORFfinder to translate the nucleic acid sequences of antibody V_H and V_L, use IMGT and VBase2 DNA PLOT to analyze their nucleic acid sequences, and determine the VDJ gene family to which they belong; mark the CDR region according to the rules of Kabat, Abm, Chothia and Contact. Then, perform BlastP with the nr library of Genbank, extract the amino acid sequence information and structural data of the antibody variable region from it to build a local antibody structure database, and divide the antibody sequences into four categories according to the antibody species and light and heavy chain types, among which the tissue sources are selected as Homo sapiens and Mus musculus.

The 200 human and mouse antibody variable region sequences with the highest similarity scores were counted to determine the percentage of amino acid types at each position; the residues with more than 95% different amino acids in the mouse alignment sequence were defined as abnormal residues (preternatural residues), and correspondingly, the residues with more than 95% different amino acids in the human alignment sequence were also defined as differential residues (differential residues); then the three-dimensional structure modeling and structural analysis of the antibody was performed, that is, the variable region homology modeling was performed using SWISS-MODEL, and the results were optimized using Energy Minimisation of Swiss-Pdb Viewer3.7 under the Gromos96 force field. The five antibody structures with the highest total similarity score with the Fv composed of antibody V_L and V_H were selected from the PDB. The interactive fit function of Swiss-Pdb Viewer 3.7 was used to spatially splice the antibody V_H and V_L with the antibody structure as a template. All results were calculated using the Force Field of the Gromos96 force field. The docking model with the minimum energy value was optimized by molecular mechanics using Discover under the Cvff force field to obtain an accurate three-dimensional model. Naccess was used to calculate the antibody V_H, V_L and their relative surface accessibility using the van der Waals radius as a parameter. FindHBond in Chimera was used to analyze the intramolecular and intermolecular hydrogen bond interactions of V_H and V_L. Finally, the candidate mutation sites were determined, and finally the humanized antibody was modeled and analyzed, and the structure was analyzed. Therefore, the humanized V_H and V_L genes obtained can encode the correct antibody variable region.

Tek Biotech has been committed to antibody research for many years. Antibody humanization is an important part of experimental research on the production and preparation of recombinant antibodies. Obtaining humanized antibodies with high specificity and affinity plays an important role in effective antibody treatment of many diseases. We have rich experience in antibody engineering construction. Based on the antibody phage display library platform, we can provide chimeric antibody modification services with high affinity and low immunogenicity.

Tek Biotech has mature antibody humanization technology, which can humanize antibody sequences and ensure that the affinity of the modified humanized antibodies remains at the same order of magnitude. We provide guaranteed humanization services to facilitate your experiments. We are committed to building a complete one-stop technical service platform, which can provide a series of services from bioinformatics analysis, humanized antibody design, antibody expression and purification, to affinity determination, antibody affinity maturation, etc. to meet the needs of different customers.