Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Novel affinity ligand polypeptide library of immunoglobulin G constructed based on protein A affinity model and application of design method

An immunoglobulin and affinity technology, applied in the field of affinity ligands, can solve the problems of poor specificity and affinity

Active Publication Date: 2015-06-24
TIANJIN UNIV
View PDF5 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the small molecule ligands discussed above have shown great advantages in antibody purification research, they also have disadvantages compared with SpA affinity media, such as poor specificity and affinity.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Novel affinity ligand polypeptide library of immunoglobulin G constructed based on protein A affinity model and application of design method
  • Novel affinity ligand polypeptide library of immunoglobulin G constructed based on protein A affinity model and application of design method
  • Novel affinity ligand polypeptide library of immunoglobulin G constructed based on protein A affinity model and application of design method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1 Obtaining a Simplified Affinity Binding Model of SpA

[0049] The absolute binding free energy of the S. aureus protein SpA and hIgG1 complex was first calculated using the MM / PBSA method. Then, a free energy decomposition method based on MM / PBSA was used to analyze the molecular mechanism of the high affinity between SpA and hIgG1 and to analyze the contribution of the residues on the binding surface of the SpA-hIgG1 complex to the binding free energy. Hotspot residues for the interaction of SpA and hIgG1 were identified based on the free energy contribution of each residue and the analysis of residue interaction pairs. Finally, an affinity binding model of SpA was constructed based on the molecular mechanism and hotspot residues obtained from the above analysis.

[0050] The SpA-hIgG1 complex molecular system was used for molecular dynamics simulations. Each complex model contains a single chain of the Fc fragment and the B domain of SpA. The model struct...

Embodiment 2

[0065] Example 2 Construction of Polypeptide Library

[0066] 1. Determine the length of the peptide sequence

[0067] A known:

[0068] The length of the peptide bond ≈

[0069] Amino acid backbone length ≈

[0070] Inserting an amino acid residue requires 2 peptide bond lengths and the main chain length of an amino acid ≈2×1.33+2.78=

[0071] Inserting two amino acid residues requires 3 peptide bond lengths and two amino acid backbone lengths ≈3×1.33+2×2.78=

[0072] image 3 An example of insertion of one amino acid (the line inside the dotted box represents the added bond). If the distance between the C and N terminals between two hotspot residues (insertion distance) , an amino acid residue can be inserted; if ≤insertion distance Then consider inserting two amino acid residues; if insertion distance> Three or more amino acid residues need to be inserted.

[0073] According to the distance between the corresponding N and C terminals of the two hotspot r...

Embodiment 3

[0086] Example 3 Docking of polypeptide molecules and Fc fragments

[0087] 1. Vina docking of polypeptide and Fc fragment

[0088] Since a single residue has a strong affinity with Fc, it does not necessarily mean that the polypeptide composed of residues has a strong interaction with Fc, so continue to use vina to sequentially dock all the polypeptides in the polypeptide library with the CBC of the Fc fragment. The docking results showed that all the peptides could bind to the Fc fragment, and the predicted binding free energy of all the peptides was between -4.5 and -8.2 kcal / mol, which was in line with the moderate affinity requirement of the affinity ligand (the binding constant was between 10 4 ~10 8 m -1 between). Among them, the peptides with binding free energy around -6.5kcal / mol were the most distributed. In order to avoid missed selection during screening, polypeptide molecules with binding free energy lower than -6.5kcal / mol were selected, and a total of 754 p...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

Disclosed in the present invention are an affinity ligand polypeptide library of immunoglobulin G constructed based on a protein A affinity model and the use thereof. On the basis of a human IgG-protein A composite structure, a key residue of protein A having a relatively high affinity effect on a human IgG is obtained by parsing, and based on which an affinity polypeptide library of IgG is constructed. Further, a candidate polypeptide is gradually screened using an amino acid positioning method, molecular docking and molecular dynamics simulation means. A polypeptide affinity ligand capable of effectively separating and purifying IgG is determined through an affinity chromatography test method.

Description

technical field [0001] The invention relates to an affinity ligand technology for designing a target protein by using molecular simulation bionics and purifying the target protein by using an affinity chromatography technology, belonging to the technical field of computer simulation and protein separation and purification in biotechnology. Background technique [0002] Antibody (immunoglobulin, Ig) is a type of glycoprotein located in animal blood and tissue fluid and produced by B lymphocytes in the immune response to antigens. The high affinity between antibody and antigen makes it widely used in biological research and clinical treatment. Especially with the gradual maturity of gene technology and cloning technology, monoclonal antibody has become an effective drug for treating inflammation, tumor and infectious diseases. At present, about 20 monoclonal antibody drugs have been approved by the FDA, and at least 300 are still under development. In 2006, the output value o...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C40B40/10C07K7/06C40B50/02C40B30/02G16B20/30G16B20/50G16B35/10G16B35/20
CPCC07K7/06G01N33/6854G16B20/00G16B35/00G16C20/60G16B20/30G16B35/10G16B35/20G16B20/50
Inventor 孙彦赵韦韦刘夫锋史清洪
Owner TIANJIN UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com