Document Detail


A biomimetic Protein G affinity adsorbent: an Ugi ligand for immunoglobulins and Fab fragments based on the third IgG-binding domain of Protein G.
MedLine Citation:
PMID:  23456743     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
This work reports the development of a synthetic affinity adsorbent for immunoglobulins based on the Fab-binding domain of Streptococcal Protein G (SpG-domain III). The ligand (A2C7I1) was synthesized by the four-component Ugi reaction to generate a substituted peptoidal scaffold mimicking key amino acid residues of SpG. Computer-aided analysis suggests a putative binding site on the CH 1 domain of the Fab molecule. In silico studies, supported by affinity chromatography in comparison with immobilized SpG, as well as analytical characterization by liquid chromatography/electrospray ionization-mass spectrometry and (1) H nuclear magnetic resonance of the ligand synthesized in solution, indicated the authenticity and suitability of the designed ligand for the purification of immunoglobulins. The immobilized ligand displayed an apparent static binding capacity of ~17 mg IgG ml(-1) and a dissociation constant of 5.34 × 10(-5)  M. Preparative chromatography demonstrated the ability of the immobilized ligand to purify IgG and Fab fragments from crude mammalian and yeast cell cultures, under near physiological ionic strength and pH, to yield proteins of 99% and 93% purity, respectively.
Authors:
Graziella El Khoury; Christopher R Lowe
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Journal of molecular recognition : JMR     Volume:  26     ISSN:  1099-1352     ISO Abbreviation:  J. Mol. Recognit.     Publication Date:  2013 Apr 
Date Detail:
Created Date:  2013-03-04     Completed Date:  2013-08-09     Revised Date:  2013-08-19    
Medline Journal Info:
Nlm Unique ID:  9004580     Medline TA:  J Mol Recognit     Country:  England    
Other Details:
Languages:  eng     Pagination:  190-200     Citation Subset:  IM    
Copyright Information:
Copyright © 2013 John Wiley & Sons, Ltd.
Affiliation:
Institute of Biotechnology, Department of Chemical Engineering and Biotechnology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QT, UK. ge235@cam.ac.uk
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MeSH Terms
Descriptor/Qualifier:
Adsorption
Amino Acid Motifs
Animals
Bacterial Proteins / chemistry*
CHO Cells
Cell Extracts / isolation & purification
Chromatography, Affinity / methods
Cricetinae
Hydrogen-Ion Concentration
Immobilized Proteins / chemical synthesis,  chemistry
Immunoglobulin Fab Fragments / chemistry,  isolation & purification*
Immunoglobulin G / chemistry,  isolation & purification*
Ligands
Molecular Docking Simulation
Molecular Mimicry
Peptide Fragments / chemical synthesis,  chemistry*
Protein Binding
Protein Interaction Domains and Motifs
Sepharose / chemistry
Solid-Phase Synthesis Techniques
Yeasts
Chemical
Reg. No./Substance:
0/Bacterial Proteins; 0/Cell Extracts; 0/IgG Fc-binding protein, Streptococcus; 0/Immobilized Proteins; 0/Immunoglobulin Fab Fragments; 0/Immunoglobulin G; 0/Ligands; 0/Peptide Fragments; 9012-36-6/Sepharose

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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